/var/spool/slurm/d/job2706798/slurm_script: line 32: fg: no job control Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_26psu_1_S13_R1_001_val_1.fq.gz 16C_26psu_1_S13_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.2/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 99 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 147 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAGGAAGTGTATGATATTAGTTTTTTTTTGTTTTTTTTTTGTTTTTTTTGTAAATGGATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP >>> Writing bisulfite mapping results to 16C_26psu_1_S13_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 80801 (80.80%) aligned concordantly 0 times 8743 (8.74%) aligned concordantly exactly 1 time 10456 (10.46%) aligned concordantly >1 times 19.20% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 80778 (80.78%) aligned concordantly 0 times 8666 (8.67%) aligned concordantly exactly 1 time 10556 (10.56%) aligned concordantly >1 times 19.22% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 82086 (82.09%) aligned concordantly 0 times 8052 (8.05%) aligned concordantly exactly 1 time 9862 (9.86%) aligned concordantly >1 times 17.91% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 82161 (82.16%) aligned concordantly 0 times 8075 (8.07%) aligned concordantly exactly 1 time 9764 (9.76%) aligned concordantly >1 times 17.84% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 36060 Mapping efficiency: 36.1% Sequence pairs with no alignments under any condition: 47402 Sequence pairs did not map uniquely: 16538 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 9306 ((converted) top strand) GA/CT/CT: 8657 (complementary to (converted) top strand) GA/CT/GA: 8718 (complementary to (converted) bottom strand) CT/GA/GA: 9379 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1365742 Total methylated C's in CpG context: 106157 Total methylated C's in CHG context: 2095 Total methylated C's in CHH context: 9506 Total methylated C's in Unknown context: 106 Total unmethylated C's in CpG context: 31364 Total unmethylated C's in CHG context: 335861 Total unmethylated C's in CHH context: 880759 Total unmethylated C's in Unknown context: 612 C methylated in CpG context: 77.2% C methylated in CHG context: 0.6% C methylated in CHH context: 1.1% C methylated in unknown context (CN or CHN): 14.8% Bismark completed in 0d 0h 0m 41s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_32psu_3_S7_R1_001_val_1.fq.gz 8C_32psu_3_S7_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.2/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 99 NC_027304.1_GA_converted 12752628 1 97M = 12752628 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 147 NC_027304.1_GA_converted 12752628 1 97M = 12752628 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA F:FFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFF:F:FFFFFFFFFFF:FFFFFFFFFFFFFFFF:FF,F:FFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 83 NC_027301.1_CT_converted 36772130 1 97M = 36772130 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 163 NC_027301.1_CT_converted 36772130 1 97M = 36772130 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 8C_32psu_3_S7_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 78324 (78.32%) aligned concordantly 0 times 9359 (9.36%) aligned concordantly exactly 1 time 12317 (12.32%) aligned concordantly >1 times 21.68% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 79743 (79.74%) aligned concordantly 0 times 8587 (8.59%) aligned concordantly exactly 1 time 11670 (11.67%) aligned concordantly >1 times 20.26% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 78322 (78.32%) aligned concordantly 0 times 9239 (9.24%) aligned concordantly exactly 1 time 12439 (12.44%) aligned concordantly >1 times 21.68% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 79734 (79.73%) aligned concordantly 0 times 8538 (8.54%) aligned concordantly exactly 1 time 11728 (11.73%) aligned concordantly >1 times 20.27% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 37892 Mapping efficiency: 37.9% Sequence pairs with no alignments under any condition: 42123 Sequence pairs did not map uniquely: 19985 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 9846 ((converted) top strand) GA/CT/CT: 9164 (complementary to (converted) top strand) GA/CT/GA: 9084 (complementary to (converted) bottom strand) CT/GA/GA: 9798 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1343247 Total methylated C's in CpG context: 106325 Total methylated C's in CHG context: 1916 Total methylated C's in CHH context: 11636 Total methylated C's in Unknown context: 96 Total unmethylated C's in CpG context: 31269 Total unmethylated C's in CHG context: 334400 Total unmethylated C's in CHH context: 857701 Total unmethylated C's in Unknown context: 600 C methylated in CpG context: 77.3% C methylated in CHG context: 0.6% C methylated in CHH context: 1.3% C methylated in unknown context (CN or CHN): 13.8% Bismark completed in 0d 0h 0m 34s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_26psu_2_S10_R1_001_val_1.fq.gz 8C_26psu_2_S10_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.2/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 99 NC_027317.1_CT_converted 34396170 0 39M = 34396170 -39 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 147 NC_027317.1_CT_converted 34396170 0 39M = 34396170 -39 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 83 NC_027317.1_GA_converted 38624794 0 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 163 NC_027317.1_GA_converted 38624794 0 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP >>> Writing bisulfite mapping results to 8C_26psu_2_S10_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 78843 (78.84%) aligned concordantly 0 times 9577 (9.58%) aligned concordantly exactly 1 time 11580 (11.58%) aligned concordantly >1 times 21.16% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 80386 (80.39%) aligned concordantly 0 times 8904 (8.90%) aligned concordantly exactly 1 time 10710 (10.71%) aligned concordantly >1 times 19.61% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 80375 (80.38%) aligned concordantly 0 times 8949 (8.95%) aligned concordantly exactly 1 time 10676 (10.68%) aligned concordantly >1 times 19.62% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 78599 (78.60%) aligned concordantly 0 times 9888 (9.89%) aligned concordantly exactly 1 time 11513 (11.51%) aligned concordantly >1 times 21.40% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 39831 Mapping efficiency: 39.8% Sequence pairs with no alignments under any condition: 42072 Sequence pairs did not map uniquely: 18097 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 10319 ((converted) top strand) GA/CT/CT: 9563 (complementary to (converted) top strand) GA/CT/GA: 9433 (complementary to (converted) bottom strand) CT/GA/GA: 10516 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1474963 Total methylated C's in CpG context: 112454 Total methylated C's in CHG context: 2337 Total methylated C's in CHH context: 9950 Total methylated C's in Unknown context: 93 Total unmethylated C's in CpG context: 32355 Total unmethylated C's in CHG context: 362797 Total unmethylated C's in CHH context: 955070 Total unmethylated C's in Unknown context: 681 C methylated in CpG context: 77.7% C methylated in CHG context: 0.6% C methylated in CHH context: 1.0% C methylated in unknown context (CN or CHN): 12.0% Bismark completed in 0d 0h 0m 32s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.2/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz (100001 sequences in total) Input files are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP >>> Writing bisulfite mapping results to CTRL_8C_26psu_1_S17_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 80332 (80.33%) aligned concordantly 0 times 9598 (9.60%) aligned concordantly exactly 1 time 10070 (10.07%) aligned concordantly >1 times 19.67% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 81256 (81.26%) aligned concordantly 0 times 9060 (9.06%) aligned concordantly exactly 1 time 100000 reads; of these:9684 ( 9.68100000% () aligned concordantly >1 times 18.74% overall alignment rate100.00 %) were paired; of these: 80483 (80.48%) aligned concordantly 0 times 9450 (9.45%) aligned concordantly exactly 1 time 10067 (10.07%) aligned concordantly >1 times 19.52% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 81232 (81.23%) aligned concordantly 0 times 9029 (9.03%) aligned concordantly exactly 1 time 9739 (9.74%) aligned concordantly >1 times 18.77% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq, CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq, CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 39628 Mapping efficiency: 39.6% Sequence pairs with no alignments under any condition: 44283 Sequence pairs did not map uniquely: 16089 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 10213 ((converted) top strand) GA/CT/CT: 9698 (complementary to (converted) top strand) GA/CT/GA: 9694 (complementary to (converted) bottom strand) CT/GA/GA: 10023 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1490664 Total methylated C's in CpG context: 105847 Total methylated C's in CHG context: 2114 Total methylated C's in CHH context: 9210 Total methylated C's in Unknown context: 96 Total unmethylated C's in CpG context: 30451 Total unmethylated C's in CHG context: 346371 Total unmethylated C's in CHH context: 996671 Total unmethylated C's in Unknown context: 695 C methylated in CpG context: 77.7% C methylated in CHG context: 0.6% C methylated in CHH context: 0.9% C methylated in unknown context (CN or CHN): 12.1% Bismark completed in 0d 0h 0m 34s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_32psu_4_S4_R1_001_val_1.fq.gz 16C_32psu_4_S4_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.2/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 16C_32psu_4_S4_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 83473 (83.47%) aligned concordantly 0 times 8499 (8.50%) aligned concordantly exactly 1 time 8028 (8.03%) aligned concordantly >1 times 16.53% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 83362 (83.36%) aligned concordantly 0 times 8563 (8.56%) aligned concordantly exactly 1 time 8075 (8.07%100000) aligned concordantly >1 times reads; of these: 16.64 %100000 overall alignment rate ( 100.00%) were paired; of these: 83548 (83.55%) aligned concordantly 0 times 8474 (8.47%) aligned concordantly exactly 1 time 7978 (7.98%) aligned concordantly >1 times 16.45% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 83578 (83.58%) aligned concordantly 0 times 8457 (8.46%) aligned concordantly exactly 1 time 7965 (7.96%) aligned concordantly >1 times 16.42% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 36743 Mapping efficiency: 36.7% Sequence pairs with no alignments under any condition: 50590 Sequence pairs did not map uniquely: 12667 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 9142 ((converted) top strand) GA/CT/CT: 9198 (complementary to (converted) top strand) GA/CT/GA: 9061 (complementary to (converted) bottom strand) CT/GA/GA: 9342 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1501518 Total methylated C's in CpG context: 105225 Total methylated C's in CHG context: 2340 Total methylated C's in CHH context: 9628 Total methylated C's in Unknown context: 117 Total unmethylated C's in CpG context: 31517 Total unmethylated C's in CHG context: 356615 Total unmethylated C's in CHH context: 996193 Total unmethylated C's in Unknown context: 769 C methylated in CpG context: 77.0% C methylated in CHG context: 0.7% C methylated in CHH context: 1.0% C methylated in unknown context (CN or CHN): 13.2% Bismark completed in 0d 0h 0m 33s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_26psu_1_S13_R1_001_val_1.fq.gz 16C_26psu_1_S13_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.3/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 99 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 147 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAGGAAGTGTATGATATTAGTTTTTTTTTGTTTTTTTTTTGTTTTTTTTGTAAATGGATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP >>> Writing bisulfite mapping results to 16C_26psu_1_S13_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 79369 (79.37%) aligned concordantly 0 times 7947 (7.95%) aligned concordantly exactly 1 time 12684 (12.68%) aligned concordantly >1 times 20.63% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 78042 (78.04%) aligned concordantly 0 times 8507 (8.51%) aligned concordantly exactly 1 time 13451 (13.45%) aligned concordantly >1 times 21.96% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 77944 (77.94%) aligned concordantly 0 times 8529 (8.53%) aligned concordantly exactly 1 time 13527 (13.53%) aligned concordantly >1 times 22.06% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 79460 (79.46%) aligned concordantly 0 times 7967 (7.97%) aligned concordantly exactly 1 time 12573 (12.57%) aligned concordantly >1 times 20.54% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 39503 Mapping efficiency: 39.5% Sequence pairs with no alignments under any condition: 42802 Sequence pairs did not map uniquely: 17695 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 10135 ((converted) top strand) GA/CT/CT: 9511 (complementary to (converted) top strand) GA/CT/GA: 9607 (complementary to (converted) bottom strand) CT/GA/GA: 10250 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1507533 Total methylated C's in CpG context: 115468 Total methylated C's in CHG context: 2586 Total methylated C's in CHH context: 11981 Total methylated C's in Unknown context: 310 Total unmethylated C's in CpG context: 34821 Total unmethylated C's in CHG context: 371395 Total unmethylated C's in CHH context: 971282 Total unmethylated C's in Unknown context: 1269 C methylated in CpG context: 76.8% C methylated in CHG context: 0.7% C methylated in CHH context: 1.2% C methylated in unknown context (CN or CHN): 19.6% Bismark completed in 0d 0h 0m 34s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_32psu_3_S7_R1_001_val_1.fq.gz 8C_32psu_3_S7_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.3/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 99 NC_027318.1_GA_converted 62868662 1 97M = 62868662 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 147 NC_027318.1_GA_converted 62868662 1 97M = 62868662 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA F:FFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFF:F:FFFFFFFFFFF:FFFFFFFFFFFFFFFF:FF,F:FFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 83 NC_027318.1_CT_converted 25087205 1 97M = 25087205 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 163 NC_027318.1_CT_converted 25087205 1 97M = 25087205 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 8C_32psu_3_S7_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 76809 (76.81%) aligned concordantly 0 times 8324 (8.32%) aligned concordantly exactly 1 time 14867 (14.87%) aligned concordantly >1 times 23.19% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 76752 (76.75%) aligned concordantly 0 times 8269 (8.27%) aligned concordantly exactly 1 time 14979 (14.98%) aligned concordantly >1 times 23.25% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75318 (75.32%) aligned concordantly 0 times 8998 (9.00%) aligned concordantly exactly 1 time 15684 (15.68%) aligned concordantly >1 times 24.68% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75272 (75.27%) aligned concordantly 0 times 9002 (9.00%) aligned concordantly exactly 1 time 15726 (15.73%) aligned concordantly >1 times 24.73% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 41092 Mapping efficiency: 41.1% Sequence pairs with no alignments under any condition: 37659 Sequence pairs did not map uniquely: 21249 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 10686 ((converted) top strand) GA/CT/CT: 9953 (complementary to (converted) top strand) GA/CT/GA: 9862 (complementary to (converted) bottom strand) CT/GA/GA: 10591 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1467653 Total methylated C's in CpG context: 115193 Total methylated C's in CHG context: 2311 Total methylated C's in CHH context: 14530 Total methylated C's in Unknown context: 279 Total unmethylated C's in CpG context: 34582 Total unmethylated C's in CHG context: 365937 Total unmethylated C's in CHH context: 935100 Total unmethylated C's in Unknown context: 1280 C methylated in CpG context: 76.9% C methylated in CHG context: 0.6% C methylated in CHH context: 1.5% C methylated in unknown context (CN or CHN): 17.9% Bismark completed in 0d 0h 0m 34s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_26psu_2_S10_R1_001_val_1.fq.gz 8C_26psu_2_S10_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.3/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 99 NC_027317.1_CT_converted 34396170 0 39M = 34396170 -39 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 147 NC_027317.1_CT_converted 34396170 0 39M = 34396170 -39 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 83 NC_027317.1_GA_converted 38624794 0 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 163 NC_027317.1_GA_converted 38624794 0 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP >>> Writing bisulfite mapping results to 8C_26psu_2_S10_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 77476 (77.48%) aligned concordantly 0 times 8778 (8.78%) aligned concordantly exactly 1 time 13746 (13.75%) aligned concordantly >1 times 22.52% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75678 (75.68%) aligned concordantly 0 times 9644 (9.64%) aligned concordantly exactly 1 time 14678 (14.68%) aligned concordantly >1 times 24.32% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 77522 (77.52%) aligned concordantly 0 times 8775 (8.78%) aligned concordantly exactly 1 time 13703 (13.70%) aligned concordantly >1 times 22.48% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75920 (75.92%) aligned concordantly 0 times 9375 (9.38%) aligned concordantly exactly 1 time 14705 (14.71%) aligned concordantly >1 times 24.08% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 43156 Mapping efficiency: 43.2% Sequence pairs with no alignments under any condition: 37605 Sequence pairs did not map uniquely: 19239 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 11132 ((converted) top strand) GA/CT/CT: 10388 (complementary to (converted) top strand) GA/CT/GA: 10286 (complementary to (converted) bottom strand) CT/GA/GA: 11350 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1607213 Total methylated C's in CpG context: 121154 Total methylated C's in CHG context: 2788 Total methylated C's in CHH context: 13592 Total methylated C's in Unknown context: 283 Total unmethylated C's in CpG context: 35601 Total unmethylated C's in CHG context: 395968 Total unmethylated C's in CHH context: 1038110 Total unmethylated C's in Unknown context: 1335 C methylated in CpG context: 77.3% C methylated in CHG context: 0.7% C methylated in CHH context: 1.3% C methylated in unknown context (CN or CHN): 17.5% Bismark completed in 0d 0h 0m 34s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.3/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz (100001 sequences in total) Input files are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP >>> Writing bisulfite mapping results to CTRL_8C_26psu_1_S17_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 77779 (77.78%) aligned concordantly 0 times 9273 (9.27%) aligned concordantly exactly 1 time 12948 (12.95%) aligned concordantly >1 times 22.22% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 77606 (77.61%) aligned concordantly 0 times 9347 (9.35%) aligned concordantly exactly 1 time 13047 (13.05%) aligned concordantly >1 times 22.39% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 78466 (78.47%) aligned concordantly 0 times 9010 (9.01%) aligned concordantly exactly 1 time 12524 (12.52%) aligned concordantly >1 times 21.53% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 78470 (78.47%) aligned concordantly 0 times 8933 (8.93%) aligned concordantly exactly 1 time 12597 (12.60%) aligned concordantly >1 times 21.53% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq, CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq, CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 43058 Mapping efficiency: 43.1% Sequence pairs with no alignments under any condition: 39777 Sequence pairs did not map uniquely: 17165 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 11063 ((converted) top strand) GA/CT/CT: 10550 (complementary to (converted) top strand) GA/CT/GA: 10608 (complementary to (converted) bottom strand) CT/GA/GA: 10837 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1629830 Total methylated C's in CpG context: 114414 Total methylated C's in CHG context: 2551 Total methylated C's in CHH context: 11165 Total methylated C's in Unknown context: 285 Total unmethylated C's in CpG context: 33820 Total unmethylated C's in CHG context: 380090 Total unmethylated C's in CHH context: 1087790 Total unmethylated C's in Unknown context: 1297 C methylated in CpG context: 77.2% C methylated in CHG context: 0.7% C methylated in CHH context: 1.0% C methylated in unknown context (CN or CHN): 18.0% Bismark completed in 0d 0h 0m 34s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_32psu_4_S4_R1_001_val_1.fq.gz 16C_32psu_4_S4_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.3/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.3 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 16C_32psu_4_S4_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz 100000 reads; of these: 100000100000 ( reads; of these: 100000 (100.00%) were paired; of these: 80967 (100.0080.97%%) were paired; of these:) aligned concordantly 0 times 808498570 ( (80.858.57%%) aligned concordantly 0 times) aligned concordantly exactly 1 time 866410463 ( (8.6610.46%%) aligned concordantly exactly 1 time) aligned concordantly >1 times 19.0310487% ( overall alignment rate10.49 %) aligned concordantly >1 times 19.15% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 80876 (80.88%) aligned concordantly 0 times 8619 (8.62%) aligned concordantly exactly 1 time 10505 (10.51%) aligned concordantly >1 times 19.12% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 81083 (81.08%) aligned concordantly 0 times 8489 (8.49%) aligned concordantly exactly 1 time 10428 (10.43%) aligned concordantly >1 times 18.92% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 40669 Mapping efficiency: 40.7% Sequence pairs with no alignments under any condition: 45723 Sequence pairs did not map uniquely: 13608 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 10049 ((converted) top strand) GA/CT/CT: 10236 (complementary to (converted) top strand) GA/CT/GA: 10104 (complementary to (converted) bottom strand) CT/GA/GA: 10280 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1671819 Total methylated C's in CpG context: 115871 Total methylated C's in CHG context: 2870 Total methylated C's in CHH context: 13994 Total methylated C's in Unknown context: 362 Total unmethylated C's in CpG context: 35330 Total unmethylated C's in CHG context: 397248 Total unmethylated C's in CHH context: 1106506 Total unmethylated C's in Unknown context: 1520 C methylated in CpG context: 76.6% C methylated in CHG context: 0.7% C methylated in CHH context: 1.2% C methylated in unknown context (CN or CHN): 19.2% Bismark completed in 0d 0h 0m 35s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_26psu_1_S13_R1_001_val_1.fq.gz 16C_26psu_1_S13_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.4/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 99 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 147 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAGGAAGTGTATGATATTAGTTTTTTTTTGTTTTTTTTTTGTTTTTTTTGTAAATGGATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP >>> Writing bisulfite mapping results to 16C_26psu_1_S13_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 77147 (77.15%) aligned concordantly 0 times 7888 (7.89%) aligned concordantly exactly 1 time 14965 (14.96%) aligned concordantly >1 times 22.85% overall alignment rate 100000100000 reads; of these: reads; of these:100000 reads; of these: 100000100000 ( (100000 (100.00100.00%100.00%) were paired; of these:%) were paired; of these: ) were paired; of these: 77287 75859 (75720 (77.29 (75.86%75.72%) aligned concordantly 0 times%) aligned concordantly 0 times ) aligned concordantly 0 times 7837 8322 (8348 (7.84 (8.32%8.35%) aligned concordantly exactly 1 time%) aligned concordantly exactly 1 time ) aligned concordantly exactly 1 time 14876 15819 (15932 (14.88 (15.82%15.93%) aligned concordantly >1 times%) aligned concordantly >1 times ) aligned concordantly >1 times 22.71 24.14%24.28% overall alignment rate% overall alignment rate overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 42211 Mapping efficiency: 42.2% Sequence pairs with no alignments under any condition: 39299 Sequence pairs did not map uniquely: 18490 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 10802 ((converted) top strand) GA/CT/CT: 10193 (complementary to (converted) top strand) GA/CT/GA: 10283 (complementary to (converted) bottom strand) CT/GA/GA: 10933 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1618025 Total methylated C's in CpG context: 122997 Total methylated C's in CHG context: 3121 Total methylated C's in CHH context: 15633 Total methylated C's in Unknown context: 611 Total unmethylated C's in CpG context: 37723 Total unmethylated C's in CHG context: 398671 Total unmethylated C's in CHH context: 1039880 Total unmethylated C's in Unknown context: 2246 C methylated in CpG context: 76.5% C methylated in CHG context: 0.8% C methylated in CHH context: 1.5% C methylated in unknown context (CN or CHN): 21.4% Bismark completed in 0d 0h 0m 34s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_32psu_3_S7_R1_001_val_1.fq.gz 8C_32psu_3_S7_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.4/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 99 NC_027309.1_GA_converted 104205143 1 97M = 104205143 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 147 NC_027309.1_GA_converted 104205143 1 97M = 104205143 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA F:FFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFF:F:FFFFFFFFFFF:FFFFFFFFFFFFFFFF:FF,F:FFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 83 NC_027314.1_CT_converted 77974989 1 97M = 77974989 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 163 NC_027314.1_CT_converted 77974989 1 97M = 77974989 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 8C_32psu_3_S7_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 74549 (74.55%) aligned concordantly 0 times 8146 (8.15%) aligned concordantly exactly 1 time 17305 (17.30%) aligned concordantly >1 times 25.45% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 73067 (73.07%) aligned concordantly 0 times 8774 (8.77%) aligned concordantly exactly 1 time 18159 (18.16%) aligned concordantly >1 times 26.93% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 73012 (73.01%) aligned concordantly 0 times 8845 (8.85%) aligned concordantly exactly 1 time 18143 (18.14%) aligned concordantly >1 times 26.99% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 74525 (74.53%) aligned concordantly 0 times 8120 (8.12%) aligned concordantly exactly 1 time 17355 (17.36%) aligned concordantly >1 times 25.48% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 43788 Mapping efficiency: 43.8% Sequence pairs with no alignments under any condition: 34233 Sequence pairs did not map uniquely: 21979 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 11355 ((converted) top strand) GA/CT/CT: 10614 (complementary to (converted) top strand) GA/CT/GA: 10523 (complementary to (converted) bottom strand) CT/GA/GA: 11296 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1569903 Total methylated C's in CpG context: 122308 Total methylated C's in CHG context: 2803 Total methylated C's in CHH context: 18215 Total methylated C's in Unknown context: 575 Total unmethylated C's in CpG context: 37551 Total unmethylated C's in CHG context: 391507 Total unmethylated C's in CHH context: 997519 Total unmethylated C's in Unknown context: 2280 C methylated in CpG context: 76.5% C methylated in CHG context: 0.7% C methylated in CHH context: 1.8% C methylated in unknown context (CN or CHN): 20.1% Bismark completed in 0d 0h 0m 36s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_26psu_2_S10_R1_001_val_1.fq.gz 8C_26psu_2_S10_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.4/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 99 NC_027317.1_CT_converted 34396170 0 39M = 34396170 -39 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 147 NC_027317.1_CT_converted 34396170 0 39M = 34396170 -39 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 83 NC_027317.1_GA_converted 38624794 0 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 163 NC_027317.1_GA_converted 38624794 0 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP >>> Writing bisulfite mapping results to 8C_26psu_2_S10_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 75389 (75.39%) aligned concordantly 0 times 8581 (8.58%) aligned concordantly exactly 1 time 16030 (16.03%) aligned concordantly >1 times 24.61% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75294 (75.29%) aligned concordantly 0 times 8640 (8.64%) aligned concordantly exactly 1 time 16066 (16.07%) aligned concordantly >1 times 24.71% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 73451 (73.45%) aligned concordantly 0 times 9459 (9.46%) aligned concordantly exactly 1 time 17090 (17.09%) aligned concordantly >1 times 26.55% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 73637 (73.64%) aligned concordantly 0 times 9183 (9.18%) aligned concordantly exactly 1 time 17180 (17.18%) aligned concordantly >1 times 26.36% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 45965 Mapping efficiency: 46.0% Sequence pairs with no alignments under any condition: 34133 Sequence pairs did not map uniquely: 19902 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 11861 ((converted) top strand) GA/CT/CT: 11125 (complementary to (converted) top strand) GA/CT/GA: 10939 (complementary to (converted) bottom strand) CT/GA/GA: 12040 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1720024 Total methylated C's in CpG context: 128223 Total methylated C's in CHG context: 3328 Total methylated C's in CHH context: 17244 Total methylated C's in Unknown context: 551 Total unmethylated C's in CpG context: 38647 Total unmethylated C's in CHG context: 423970 Total unmethylated C's in CHH context: 1108612 Total unmethylated C's in Unknown context: 2372 C methylated in CpG context: 76.8% C methylated in CHG context: 0.8% C methylated in CHH context: 1.5% C methylated in unknown context (CN or CHN): 18.9% Bismark completed in 0d 0h 0m 34s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.4/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz (100001 sequences in total) Input files are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP >>> Writing bisulfite mapping results to CTRL_8C_26psu_1_S17_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 76392 (76.39%) aligned concordantly 0 times 8872 (8.87%) aligned concordantly exactly 1 time 14736 (14.74%) aligned concordantly >1 times 23.61% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 76357 (76.36%) aligned concordantly 0 times 8837 (8.84%) aligned concordantly exactly 1 time 14806 (14.81%) aligned concordantly >1 times 23.64% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75633 (75.63%) aligned concordantly 0 times 9123 (9.12%) aligned concordantly exactly 1 time 15244 (15.24%) aligned concordantly >1 times 24.37% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75532 (75.53%) aligned concordantly 0 times 9185 (9.19%) aligned concordantly exactly 1 time 15283 (15.28%) aligned concordantly >1 times 24.47% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq, CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq, CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 45830 Mapping efficiency: 45.8% Sequence pairs with no alignments under any condition: 36362 Sequence pairs did not map uniquely: 17808 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 11759 ((converted) top strand) GA/CT/CT: 11239 (complementary to (converted) top strand) GA/CT/GA: 11296 (complementary to (converted) bottom strand) CT/GA/GA: 11536 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1742550 Total methylated C's in CpG context: 121434 Total methylated C's in CHG context: 2977 Total methylated C's in CHH context: 14216 Total methylated C's in Unknown context: 575 Total unmethylated C's in CpG context: 36466 Total unmethylated C's in CHG context: 407345 Total unmethylated C's in CHH context: 1160112 Total unmethylated C's in Unknown context: 2250 C methylated in CpG context: 76.9% C methylated in CHG context: 0.7% C methylated in CHH context: 1.2% C methylated in unknown context (CN or CHN): 20.4% Bismark completed in 0d 0h 0m 36s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_32psu_4_S4_R1_001_val_1.fq.gz 16C_32psu_4_S4_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.4/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.4 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 16C_32psu_4_S4_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 79056 (79.06%) aligned concordantly 0 times 8389 (8.39%) aligned concordantly exactly 1 time 12555 (12.55%) aligned concordantly >1 times 20.94% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 78899 (78.90%) aligned concordantly 0 times 8483 (8.48%) aligned concordantly exactly 1 time 12618 (12.62%) aligned concordantly >1 times 21.10% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 78744 (78.74%) aligned concordantly 0 times 8677 (8.68%) aligned concordantly exactly 1 time 12579 (12.58%) aligned concordantly >1 times 21.26% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 78843 (78.84%) aligned concordantly 0 times 8555 (8.55%) aligned concordantly exactly 1 time 12602 (12.60%) aligned concordantly >1 times 21.16% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 43788 Mapping efficiency: 43.8% Sequence pairs with no alignments under any condition: 42040 Sequence pairs did not map uniquely: 14172 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 10825 ((converted) top strand) GA/CT/CT: 11071 (complementary to (converted) top strand) GA/CT/GA: 10919 (complementary to (converted) bottom strand) CT/GA/GA: 10973 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1806051 Total methylated C's in CpG context: 123895 Total methylated C's in CHG context: 3384 Total methylated C's in CHH context: 17873 Total methylated C's in Unknown context: 736 Total unmethylated C's in CpG context: 38542 Total unmethylated C's in CHG context: 429848 Total unmethylated C's in CHH context: 1192509 Total unmethylated C's in Unknown context: 2647 C methylated in CpG context: 76.3% C methylated in CHG context: 0.8% C methylated in CHH context: 1.5% C methylated in unknown context (CN or CHN): 21.8% Bismark completed in 0d 0h 0m 36s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_26psu_1_S13_R1_001_val_1.fq.gz 16C_26psu_1_S13_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.5/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 99 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 147 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAGGAAGTGTATGATATTAGTTTTTTTTTGTTTTTTTTTTGTTTTTTTTGTAAATGGATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP >>> Writing bisulfite mapping results to 16C_26psu_1_S13_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 73753 (73.75%) aligned concordantly 0 times 8244 (8.24%) aligned concordantly exactly 1 time 18003 (18.00%) aligned concordantly >1 times 26.25% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 73659 (73.66%) aligned concordantly 0 times 8300 (8.30%) aligned concordantly exactly 1 time 18041 (18.04%) aligned concordantly >1 times 26.34% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75004 (75.00%) aligned concordantly 0 times 7810 (7.81%) aligned concordantly exactly 1 time 17186 (17.19%) aligned concordantly >1 times 25.00% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75098 (75.10%) aligned concordantly 0 times 7860 (7.86%) aligned concordantly exactly 1 time 17042 (17.04%) aligned concordantly >1 times 24.90% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 44923 Mapping efficiency: 44.9% Sequence pairs with no alignments under any condition: 36063 Sequence pairs did not map uniquely: 19014 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 11482 ((converted) top strand) GA/CT/CT: 10878 (complementary to (converted) top strand) GA/CT/GA: 10978 (complementary to (converted) bottom strand) CT/GA/GA: 11585 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1730056 Total methylated C's in CpG context: 130764 Total methylated C's in CHG context: 3750 Total methylated C's in CHH context: 21603 Total methylated C's in Unknown context: 1090 Total unmethylated C's in CpG context: 40925 Total unmethylated C's in CHG context: 425680 Total unmethylated C's in CHH context: 1107334 Total unmethylated C's in Unknown context: 3645 C methylated in CpG context: 76.2% C methylated in CHG context: 0.9% C methylated in CHH context: 1.9% C methylated in unknown context (CN or CHN): 23.0% Bismark completed in 0d 0h 0m 36s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_32psu_3_S7_R1_001_val_1.fq.gz 8C_32psu_3_S7_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.5/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 99 NC_027312.1_GA_converted 43816323 1 97M = 43816323 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 147 NC_027312.1_GA_converted 43816323 1 97M = 43816323 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA F:FFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFF:F:FFFFFFFFFFF:FFFFFFFFFFFFFFFF:FF,F:FFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 83 NC_027321.1_CT_converted 46198681 1 97M = 46198681 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 163 NC_027321.1_CT_converted 46198681 1 97M = 46198681 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 8C_32psu_3_S7_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 72371 (72.37%) aligned concordantly 0 times 8116 (8.12%) aligned concordantly exactly 1 time 19513 (19.51%) aligned concordantly >1 times 27.63% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 70827 (70.83%) aligned concordantly 0 times 8577 (8.58%) aligned concordantly exactly 1 time 20596 (20.60%) aligned concordantly >1 times 29.17% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 70816 (70.82%) aligned concordantly 0 times 8658 (8.66%) aligned concordantly exactly 1 time 20526 (20.53%) aligned concordantly >1 times 29.18% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 72313 (72.31%) aligned concordantly 0 times 8131 (8.13%) aligned concordantly exactly 1 time 19556 (19.56%) aligned concordantly >1 times 27.69% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 46195 Mapping efficiency: 46.2% Sequence pairs with no alignments under any condition: 31191 Sequence pairs did not map uniquely: 22614 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 11946 ((converted) top strand) GA/CT/CT: 11211 (complementary to (converted) top strand) GA/CT/GA: 11132 (complementary to (converted) bottom strand) CT/GA/GA: 11906 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1666250 Total methylated C's in CpG context: 128971 Total methylated C's in CHG context: 3367 Total methylated C's in CHH context: 23962 Total methylated C's in Unknown context: 1020 Total unmethylated C's in CpG context: 40527 Total unmethylated C's in CHG context: 414718 Total unmethylated C's in CHH context: 1054705 Total unmethylated C's in Unknown context: 3579 C methylated in CpG context: 76.1% C methylated in CHG context: 0.8% C methylated in CHH context: 2.2% C methylated in unknown context (CN or CHN): 22.2% Bismark completed in 0d 0h 0m 35s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_26psu_2_S10_R1_001_val_1.fq.gz 8C_26psu_2_S10_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.5/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 99 NC_027317.1_CT_converted 34396730 1 39M = 34397170 479 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 147 NC_027317.1_CT_converted 34397170 1 39M = 34396730 -479 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 83 NC_027317.1_GA_converted 38624794 1 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 163 NC_027317.1_GA_converted 38624794 1 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP >>> Writing bisulfite mapping results to 8C_26psu_2_S10_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 73350 (73.35%) aligned concordantly 0 times 8531 (8.53%) aligned concordantly exactly 1 time 18119 (18.12%) aligned concordantly >1 times 26.65% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 73218 (73.22%) aligned concordantly 0 times 8616 (8.62%) aligned concordantly exactly 1 time 18166 (18.17%) aligned concordantly >1 times 26.78% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 71329 (71.33%) aligned concordantly 0 times 9283 (9.28%) aligned concordantly exactly 1 time 19388 (19.39%) aligned concordantly >1 times 28.67% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 71464 (71.46%) aligned concordantly 0 times 9020 (9.02%) aligned concordantly exactly 1 time 19516 (19.52%) aligned concordantly >1 times 28.54% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 48413 Mapping efficiency: 48.4% Sequence pairs with no alignments under any condition: 31106 Sequence pairs did not map uniquely: 20481 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 12494 ((converted) top strand) GA/CT/CT: 11755 (complementary to (converted) top strand) GA/CT/GA: 11518 (complementary to (converted) bottom strand) CT/GA/GA: 12646 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1820121 Total methylated C's in CpG context: 134838 Total methylated C's in CHG context: 3819 Total methylated C's in CHH context: 21511 Total methylated C's in Unknown context: 992 Total unmethylated C's in CpG context: 41434 Total unmethylated C's in CHG context: 448107 Total unmethylated C's in CHH context: 1170412 Total unmethylated C's in Unknown context: 3769 C methylated in CpG context: 76.5% C methylated in CHG context: 0.8% C methylated in CHH context: 1.8% C methylated in unknown context (CN or CHN): 20.8% Bismark completed in 0d 0h 0m 36s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.5/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz (100001 sequences in total) Input files are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP >>> Writing bisulfite mapping results to CTRL_8C_26psu_1_S17_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 73631 (73.63%) aligned concordantly 0 times 8937 (8.94%) aligned concordantly exactly 1 time 17432 (17.43%) aligned concordantly >1 times 26.37% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 73566 (73.57%) aligned concordantly 0 times 9056 (9.06%) aligned concordantly exactly 1 time 17378 (17.38%) aligned concordantly >1 times 26.43% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 74302 (74.30%) aligned concordantly 0 times 8904 (8.90%) aligned concordantly exactly 1 time 16794 (16.79%) aligned concordantly >1 times 25.70% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 74287 (74.29%) aligned concordantly 0 times 8752 (8.75%) aligned concordantly exactly 1 time 16961 (16.96%) aligned concordantly >1 times 25.71% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq, CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq, CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 48376 Mapping efficiency: 48.4% Sequence pairs with no alignments under any condition: 33354 Sequence pairs did not map uniquely: 18270 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 12413 ((converted) top strand) GA/CT/CT: 11860 (complementary to (converted) top strand) GA/CT/GA: 11977 (complementary to (converted) bottom strand) CT/GA/GA: 12126 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1846766 Total methylated C's in CpG context: 127238 Total methylated C's in CHG context: 3495 Total methylated C's in CHH context: 18539 Total methylated C's in Unknown context: 917 Total unmethylated C's in CpG context: 39022 Total unmethylated C's in CHG context: 432122 Total unmethylated C's in CHH context: 1226350 Total unmethylated C's in Unknown context: 3627 C methylated in CpG context: 76.5% C methylated in CHG context: 0.8% C methylated in CHH context: 1.5% C methylated in unknown context (CN or CHN): 20.2% Bismark completed in 0d 0h 0m 37s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_32psu_4_S4_R1_001_val_1.fq.gz 16C_32psu_4_S4_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.5/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.5 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 16C_32psu_4_S4_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 76951 (76.95%) aligned concordantly 0 times 8437 (8.44%) aligned concordantly exactly 1 time 14612 (14.61%) aligned concordantly >1 times 23.05% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 76712 (76.71%) aligned concordantly 0 times 8621 (8.62%) aligned concordantly exactly 1 time 14667 (14.67%) aligned concordantly >1 times 23.29% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 76831 (76.83%) aligned concordantly 0 times 8569 (8.57%) aligned concordantly exactly 1 time 14600 (14.60%) aligned concordantly >1 times 23.17% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 77108 (77.11%) aligned concordantly 0 times 8335 (8.34%) aligned concordantly exactly 1 time 14557 (14.56%) aligned concordantly >1 times 22.89% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 46637 Mapping efficiency: 46.6% Sequence pairs with no alignments under any condition: 38704 Sequence pairs did not map uniquely: 14659 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 11490 ((converted) top strand) GA/CT/CT: 11841 (complementary to (converted) top strand) GA/CT/GA: 11677 (complementary to (converted) bottom strand) CT/GA/GA: 11629 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1928038 Total methylated C's in CpG context: 131370 Total methylated C's in CHG context: 3927 Total methylated C's in CHH context: 21892 Total methylated C's in Unknown context: 1151 Total unmethylated C's in CpG context: 41560 Total unmethylated C's in CHG context: 458962 Total unmethylated C's in CHH context: 1270327 Total unmethylated C's in Unknown context: 4157 C methylated in CpG context: 76.0% C methylated in CHG context: 0.8% C methylated in CHH context: 1.7% C methylated in unknown context (CN or CHN): 21.7% Bismark completed in 0d 0h 0m 35s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_26psu_1_S13_R1_001_val_1.fq.gz 16C_26psu_1_S13_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.6/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R1_001_val_1.fq.gz to 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_26psu_1_S13_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_26psu_1_S13_R2_001_val_2.fq.gz to 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_26psu_1_S13_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 99 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 147 NC_027306.1_GA_converted 27312195 42 61M = 27312195 -61 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFF AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:61 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATCCATTTACAAAAAAAACAAAAAAAAAACAAAAAAAAACTAATATCATACACTTCCTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAGGAAGTGTATGATATTAGTTTTTTTTTGTTTTTTTTTTGTTTTTTTTGTAAATGGATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1262:1016_1:N:0:NGTCAA/1 77 * 0 0 * * 0 0 AATTTATTTATAAAAAAAATAAAAAAAAAATAAAAAAAAATTAATATTATATATTTTTTTT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFF:FFFFFFFFF:FFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1262:1016_2:N:0:NGTCAA/2 141 * 0 0 * * 0 0 AAAAAAAATATATAATATTAATTTTTTTTTATTTTTTTTTTATTTTTTTTATAAATAAATT FFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFF YT:Z:UP >>> Writing bisulfite mapping results to 16C_26psu_1_S13_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_26psu_1_S13_R1_001_val_1.fq.gz and 16C_26psu_1_S13_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 71593 (71.59%) aligned concordantly 0 times 8387 (8.39%) aligned concordantly exactly 1 time 20020 (20.02%) aligned concordantly >1 times 28.41% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 71617 (71.62%) aligned concordantly 0 times 8325 (8.32%) aligned concordantly exactly 1 time 20058 (20.06%) aligned concordantly >1 times 28.38% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 73003 (73.00%) aligned concordantly 0 times 7833 (7.83%) aligned concordantly exactly 1 time 19164 (19.16%) aligned concordantly >1 times 27.00% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 72960 (72.96%) aligned concordantly 0 times 7851 (7.85%) aligned concordantly exactly 1 time 19189 (19.19%) aligned concordantly >1 times 27.04% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_26psu_1_S13_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_26psu_1_S13_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_26psu_1_S13_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_26psu_1_S13_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 47338 Mapping efficiency: 47.3% Sequence pairs with no alignments under any condition: 33103 Sequence pairs did not map uniquely: 19559 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 12126 ((converted) top strand) GA/CT/CT: 11474 (complementary to (converted) top strand) GA/CT/GA: 11529 (complementary to (converted) bottom strand) CT/GA/GA: 12209 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1830499 Total methylated C's in CpG context: 137676 Total methylated C's in CHG context: 4336 Total methylated C's in CHH context: 27122 Total methylated C's in Unknown context: 1611 Total unmethylated C's in CpG context: 44056 Total unmethylated C's in CHG context: 449256 Total unmethylated C's in CHH context: 1168053 Total unmethylated C's in Unknown context: 5501 C methylated in CpG context: 75.8% C methylated in CHG context: 1.0% C methylated in CHH context: 2.3% C methylated in unknown context (CN or CHN): 22.7% Bismark completed in 0d 0h 0m 36s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_32psu_3_S7_R1_001_val_1.fq.gz 8C_32psu_3_S7_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.6/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R1_001_val_1.fq.gz to 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_32psu_3_S7_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_32psu_3_S7_R2_001_val_2.fq.gz to 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_32psu_3_S7_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 99 NC_027315.1_GA_converted 58913704 1 97M = 58913704 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 147 NC_027315.1_GA_converted 58913704 1 97M = 58913704 -97 TTTAAAAATAACCTTACTAAAAAATCACACTTTACAATAATCTAAACACTACACCCAAAAAAATACACATTACAATACAAACTAAACATCATATTAA F:FFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFF:F:FFFFFFFFFFF:FFFFFFFFFFFFFFFF:FF,F:FFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 83 NC_027317.1_CT_converted 867529 1 97M = 867529 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 163 NC_027317.1_CT_converted 867529 1 97M = 867529 -97 TTAATATGATGTTTAGTTTGTATTGTAATGTGTATTTTTTTGGGTGTAGTGTTTAGATTATTGTAAAGTGTGATTTTTTAGTAAGGTTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F AS:i:0 XS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:97 YS:i:0 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1931:1016_1:N:0:NAGATC/1 77 * 0 0 * * 0 0 TTTAAAAATAATTTTATTAAAAAATTATATTTTATAATAATTTAAATATTATGTTTAAAAAAATATGTGTTATGATATAAATTAAATGTTATATTAA FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1931:1016_2:N:0:NAGATC/2 141 * 0 0 * * 0 0 TTAATATAACATTTAATTTATATCATAACACATATTTTTTTAAACATAATATTTAAATTATTATAAAATATAATTTTTTAATAAAATTATTTTTAAA FFFFFFFF:F,FF:FFFFFFFFFFFFFFFF:FFFFFFFFFFF:F:FFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 8C_32psu_3_S7_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_32psu_3_S7_R1_001_val_1.fq.gz and 8C_32psu_3_S7_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 68677 (68.68%) aligned concordantly 0 times 8529 (8.53%) aligned concordantly exactly 1 time 22794 (22.79%) aligned concordantly >1 times 31.32% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 70111 (70.11%) aligned concordantly 0 times 8240 (8.24%) aligned concordantly exactly 1 time 21649 (21.65%) aligned concordantly >1 times 29.89% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 68706 (68.71%) aligned concordantly 0 times 8627 (8.63%) aligned concordantly exactly 1 time 22667 (22.67%) aligned concordantly >1 times 31.29% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 70056 (70.06%) aligned concordantly 0 times 8215 (8.21%) aligned concordantly exactly 1 time 21729 (21.73%) aligned concordantly >1 times 29.94% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_32psu_3_S7_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_32psu_3_S7_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_32psu_3_S7_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_32psu_3_S7_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 48514 Mapping efficiency: 48.5% Sequence pairs with no alignments under any condition: 28438 Sequence pairs did not map uniquely: 23048 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 12526 ((converted) top strand) GA/CT/CT: 11813 (complementary to (converted) top strand) GA/CT/GA: 11679 (complementary to (converted) bottom strand) CT/GA/GA: 12496 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1758005 Total methylated C's in CpG context: 135028 Total methylated C's in CHG context: 3979 Total methylated C's in CHH context: 30362 Total methylated C's in Unknown context: 1603 Total unmethylated C's in CpG context: 43343 Total unmethylated C's in CHG context: 436061 Total unmethylated C's in CHH context: 1109232 Total unmethylated C's in Unknown context: 5412 C methylated in CpG context: 75.7% C methylated in CHG context: 0.9% C methylated in CHH context: 2.7% C methylated in unknown context (CN or CHN): 22.9% Bismark completed in 0d 0h 0m 36s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 8C_26psu_2_S10_R1_001_val_1.fq.gz 8C_26psu_2_S10_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.6/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R1_001_val_1.fq.gz to 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 8C_26psu_2_S10_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 8C_26psu_2_S10_R2_001_val_2.fq.gz to 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 8C_26psu_2_S10_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 99 NC_027317.1_CT_converted 34396730 1 39M = 34397170 479 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 147 NC_027317.1_CT_converted 34397170 1 39M = 34396730 -479 TTATTGTGTGAAAGAAGGTGTGTTTTTAGTTTTTTATAG FFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:11T27 YS:i:-6 YT:Z:CP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 77 * 0 0 * * 0 0 TTATTATATAAAAAAAAATATATTTTTAATTTTTTATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 141 * 0 0 * * 0 0 TTATAAAAAATTAAAAATATATTTTTTTTTATATAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1588:1016_1:N:0:NAGCTT/1 83 NC_027317.1_GA_converted 38624794 1 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP A00177:85:HCW7VDRXX:1:2101:1588:1016_2:N:0:NAGCTT/2 163 NC_027317.1_GA_converted 38624794 1 39M = 38624794 -39 CTATAAAAAACTAAAAACACACCTTCTTTCACACAATAA FFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF AS:i:-6 XS:i:-6 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:27A11 YS:i:-6 YT:Z:CP >>> Writing bisulfite mapping results to 8C_26psu_2_S10_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 8C_26psu_2_S10_R1_001_val_1.fq.gz and 8C_26psu_2_S10_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 71259 (71.26%) aligned concordantly 0 times 8545 (8.54%) aligned concordantly exactly 1 time 20196 (20.20%) aligned concordantly >1 times 28.74% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 71140 (71.14%) aligned concordantly 0 times 8635 (8.63%) aligned concordantly exactly 1 time 20225 (20.23%) aligned concordantly >1 times 28.86% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 69333 (69.33%) aligned concordantly 0 times 9026 (9.03%) aligned concordantly exactly 1 time 21641 (21.64%) aligned concordantly >1 times 30.67% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 69214 (69.21%) aligned concordantly 0 times 9226 (9.23%) aligned concordantly exactly 1 time 21560 (21.56%) aligned concordantly >1 times 30.79% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 8C_26psu_2_S10_R1_001_val_1.fq.gz_C_to_T.fastq, 8C_26psu_2_S10_R1_001_val_1.fq.gz_G_to_A.fastq, 8C_26psu_2_S10_R2_001_val_2.fq.gz_C_to_T.fastq and 8C_26psu_2_S10_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 50709 Mapping efficiency: 50.7% Sequence pairs with no alignments under any condition: 28472 Sequence pairs did not map uniquely: 20819 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 13113 ((converted) top strand) GA/CT/CT: 12343 (complementary to (converted) top strand) GA/CT/GA: 12034 (complementary to (converted) bottom strand) CT/GA/GA: 13219 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1914285 Total methylated C's in CpG context: 140902 Total methylated C's in CHG context: 4369 Total methylated C's in CHH context: 26209 Total methylated C's in Unknown context: 1458 Total unmethylated C's in CpG context: 44152 Total unmethylated C's in CHG context: 470776 Total unmethylated C's in CHH context: 1227877 Total unmethylated C's in Unknown context: 5363 C methylated in CpG context: 76.1% C methylated in CHG context: 0.9% C methylated in CHH context: 2.1% C methylated in unknown context (CN or CHN): 21.4% Bismark completed in 0d 0h 0m 36s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.6/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz to CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz to CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz (100001 sequences in total) Input files are CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 AATAATATTTATTTATTATAAAAATAAAAATCATAAAAATTAATATAAAAATCATAAATTAAATAAAAAAAAAAAAAATAAAAAAAAATAAAAAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 TTTTTTTATTTTTTTTTATTTTTTTTTTTTTTATTTAATTTATGATTTTTATATTAATTTTTATGATTTTTATTTTTATAATAAATAAATATTATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:1298:1016_1:N:0:NTAGAG/1 77 * 0 0 * * 0 0 GATAATGTTTATTTATTGTAGAGATAGAGATTGTAGAGGTTAGTATAAAGGTTGTGAATTAGGTAGAGAGAGAGAGAGTAAGGGAGAGTGAGAGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF YT:Z:UP A00177:85:HCW7VDRXX:1:2101:1298:1016_2:N:0:NTAGAG/2 141 * 0 0 * * 0 0 CTCTCTCACTCTCCCTTACTCTCTCTCTCTCTACCTAATTCACAACCTTTATACTAACCTCTACAATCTCTATCTCTACAATAAATAAACATTATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFF:FFF:F:FFFF:FFFFFFFFFFFFFF YT:Z:UP >>> Writing bisulfite mapping results to CTRL_8C_26psu_1_S17_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 72216 (72.22%) aligned concordantly 0 times 8861 (8.86%) aligned concordantly exactly 1 time 18923 (18.92%) aligned concordantly >1 times 27.78% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 71616 (71.62%) aligned concordantly 0 times 8943 (8.94%) aligned concordantly exactly 1 time 19441 (19.44%) aligned concordantly >1 times 28.38% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 71524 (71.52%) aligned concordantly 0 times 9062 (9.06%) aligned concordantly exactly 1 time 19414 (19.41%) aligned concordantly >1 times 28.48% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 72192 (72.19%) aligned concordantly 0 times 8918 (8.92%) aligned concordantly exactly 1 time 18890 (18.89%) aligned concordantly >1 times 27.81% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_C_to_T.fastq, CTRL_8C_26psu_1_S17_R1_001_val_1.fq.gz_G_to_A.fastq, CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_C_to_T.fastq and CTRL_8C_26psu_1_S17_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 50628 Mapping efficiency: 50.6% Sequence pairs with no alignments under any condition: 30726 Sequence pairs did not map uniquely: 18646 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 12963 ((converted) top strand) GA/CT/CT: 12443 (complementary to (converted) top strand) GA/CT/GA: 12537 (complementary to (converted) bottom strand) CT/GA/GA: 12685 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 1940689 Total methylated C's in CpG context: 132803 Total methylated C's in CHG context: 3999 Total methylated C's in CHH context: 24087 Total methylated C's in Unknown context: 1387 Total unmethylated C's in CpG context: 41489 Total unmethylated C's in CHG context: 453629 Total unmethylated C's in CHH context: 1284682 Total unmethylated C's in Unknown context: 5281 C methylated in CpG context: 76.2% C methylated in CHG context: 0.9% C methylated in CHH context: 1.8% C methylated in unknown context (CN or CHN): 20.8% Bismark completed in 0d 0h 0m 38s ==================== Bismark run complete ==================== Bowtie seems to be working fine (tested command '/gscratch/srlab/programs/bowtie2-2.3.4.1-linux-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools path provided as: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ (absolute path is '/gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/)' FastQ format assumed (by default) Processing sequences up to read no. 100000 from the input file Each Bowtie 2 instance is going to be run with 4 threads. Please monitor performance closely and tune down if needed! Input files to be analysed (in current folder '/gscratch/scrubbed/strigg/analyses/20200612'): 16C_32psu_4_S4_R1_001_val_1.fq.gz 16C_32psu_4_S4_R2_001_val_2.fq.gz Library was specified to be not strand-specific (non-directional), therefore alignments to all four possible bisulfite strands (OT, CTOT, OB and CTOB) will be reported Output will be written into the directory: /gscratch/scrubbed/strigg/analyses/20200612/AS_0.6/ Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/strigg/analyses/20200612 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ chr NC_027300.1 (159038749 bp) chr NC_027301.1 (72943711 bp) chr NC_027302.1 (92503428 bp) chr NC_027303.1 (82398023 bp) chr NC_027304.1 (80503876 bp) chr NC_027305.1 (87043187 bp) chr NC_027306.1 (58785265 bp) chr NC_027307.1 (26434011 bp) chr NC_027308.1 (141712163 bp) chr NC_027309.1 (116138521 bp) chr NC_027310.1 (93888508 bp) chr NC_027311.1 (91880962 bp) chr NC_027312.1 (107758822 bp) chr NC_027313.1 (93901823 bp) chr NC_027314.1 (103963436 bp) chr NC_027315.1 (87796322 bp) chr NC_027316.1 (57682537 bp) chr NC_027317.1 (70695409 bp) chr NC_027318.1 (82978132 bp) chr NC_027319.1 (86795997 bp) chr NC_027320.1 (58021487 bp) chr NC_027321.1 (63420196 bp) chr NC_027322.1 (49854004 bp) chr NC_027323.1 (48650976 bp) chr NC_027324.1 (51481326 bp) chr NC_027325.1 (47900953 bp) chr NC_027326.1 (43943985 bp) chr NC_027327.1 (39600944 bp) chr NC_027328.1 (42488238 bp) chr NC_001960.1 (16665 bp) Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz Input files are in FastQ format Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R1_001_val_1.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R1_001_val_1.fq.gz to 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R1_001_val_1.fq.gz (100001 sequences in total) Processing reads up to sequence no. 100000 from 16C_32psu_4_S4_R2_001_val_2.fq.gz Writing a C -> T converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq Writing a G -> A converted version of the input file 16C_32psu_4_S4_R2_001_val_2.fq.gz to 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file 16C_32psu_4_S4_R2_001_val_2.fq.gz (100001 sequences in total) Input files are 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/strigg/data/Ssalar/GENOMES/chr1-29MT/ with the specified options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for GAread1CTread2CTgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 AATTATCAAAATTTTTTTTTTAATTTTATTTTAAAAAAAATATTTTTATTAAAATAATAATTTAAAATATTATATTATAATAATTAAA FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 TTTAATTATTATAATATAATATTTTAAATTATTATTTTAATAAAAATATTTTTTTTAAAATAAAATTAAAAAAAAAATTTTGATAATT FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP Now starting a Bowtie 2 paired-end alignment for CTread1GAread2GAgenome (reading in sequences from 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --nofw)) Found first alignment: A00177:85:HCW7VDRXX:1:2101:2022:1016_1:N:0:NGACCA/1 77 * 0 0 * * 0 0 GATTATTGAGATTTTTTTTTTAATTTTATTTTGAAAGAGGTGTTTTTATTGGAGTAATGGTTTAAGGTATTGTATTGTAGTGGTTGAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFF:FFFF,FFFFFFFFF:F YT:Z:UP A00177:85:HCW7VDRXX:1:2101:2022:1016_2:N:0:NGACCA/2 141 * 0 0 * * 0 0 CTCAACCACTACAATACAATACCTTAAACCATTACTCCAATAAAAACACCTCTTTCAAAATAAAATTAAAAAAAAAATCTCAATAATC FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:F YT:Z:UP >>> Writing bisulfite mapping results to 16C_32psu_4_S4_R1_001_val_1_bismark_bt2_pe.bam <<< Reading in the sequence files 16C_32psu_4_S4_R1_001_val_1.fq.gz and 16C_32psu_4_S4_R2_001_val_2.fq.gz 100000 reads; of these: 100000 (100.00%) were paired; of these: 75121 (75.12%) aligned concordantly 0 times 8360 (8.36%) aligned concordantly exactly 1 time 16519 (16.52%) aligned concordantly >1 times 24.88% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 74676 (74.68%) aligned concordantly 0 times 8684 (8.68%) aligned concordantly exactly 1 time 16640 (16.64%) aligned concordantly >1 times 25.32% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 74720 (74.72%) aligned concordantly 0 times 8670 (8.67%) aligned concordantly exactly 1 time 16610 (16.61%) aligned concordantly >1 times 25.28% overall alignment rate 100000 reads; of these: 100000 (100.00%) were paired; of these: 75030 (75.03%) aligned concordantly 0 times 8468 (8.47%) aligned concordantly exactly 1 time 16502 (16.50%) aligned concordantly >1 times 24.97% overall alignment rate Processed 100000 sequences in total Successfully deleted the temporary files 16C_32psu_4_S4_R1_001_val_1.fq.gz_C_to_T.fastq, 16C_32psu_4_S4_R1_001_val_1.fq.gz_G_to_A.fastq, 16C_32psu_4_S4_R2_001_val_2.fq.gz_C_to_T.fastq and 16C_32psu_4_S4_R2_001_val_2.fq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 100000 Number of paired-end alignments with a unique best hit: 49269 Mapping efficiency: 49.3% Sequence pairs with no alignments under any condition: 35632 Sequence pairs did not map uniquely: 15099 Sequence pairs which were discarded because genomic sequence could not be extracted: 0 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 12146 ((converted) top strand) GA/CT/CT: 12511 (complementary to (converted) top strand) GA/CT/GA: 12351 (complementary to (converted) bottom strand) CT/GA/GA: 12261 ((converted) bottom strand) Final Cytosine Methylation Report ================================= Total number of C's analysed: 2041112 Total methylated C's in CpG context: 138394 Total methylated C's in CHG context: 4493 Total methylated C's in CHH context: 27719 Total methylated C's in Unknown context: 1693 Total unmethylated C's in CpG context: 44636 Total unmethylated C's in CHG context: 484479 Total unmethylated C's in CHH context: 1341391 Total unmethylated C's in Unknown context: 6242 C methylated in CpG context: 75.6% C methylated in CHG context: 0.9% C methylated in CHH context: 2.0% C methylated in unknown context (CN or CHN): 21.3% Bismark completed in 0d 0h 0m 36s ==================== Bismark run complete ====================