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 found here: '/gscratch/srlab/programs/samtools-1.9/samtools' Reference genome folder provided is /gscratch/srlab/sam/data/C_virginica/genomes/GCF_002022765.2_C_virginica-3.0/ (absolute path is '/gscratch/srlab/sam/data/C_virginica/genomes/GCF_002022765.2_C_virginica-3.0/)' Processing sequences up to read no. 27591427 from the input file Attention: using more than 4 cores per alignment thread has been reported to have diminishing returns. If possible try to limit -p to a value of 4 Input files to be analysed (in current folder '/gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/avg_reads_bismark'): /gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/cvir_bsseq_all_pe_R1.fastq.gz /gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/cvir_bsseq_all_pe_R2.fastq.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 Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/avg_reads_bismark Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/sam/data/C_virginica/genomes/GCF_002022765.2_C_virginica-3.0/ Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are /gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/cvir_bsseq_all_pe_R1.fastq.gz and /gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/cvir_bsseq_all_pe_R2.fastq.gz Input files are in FastQ format Processing reads up to sequence no. 27591427 from /gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/cvir_bsseq_all_pe_R1.fastq.gz Writing a C -> T converted version of the input file cvir_bsseq_all_pe_R1.fastq.gz to cvir_bsseq_all_pe_R1.fastq.gz_C_to_T.fastq Writing a G -> A converted version of the input file cvir_bsseq_all_pe_R1.fastq.gz to cvir_bsseq_all_pe_R1.fastq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file cvir_bsseq_all_pe_R1.fastq.gz (27591428 sequences in total) Processing reads up to sequence no. 27591427 from /gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/cvir_bsseq_all_pe_R2.fastq.gz Writing a C -> T converted version of the input file cvir_bsseq_all_pe_R2.fastq.gz to cvir_bsseq_all_pe_R2.fastq.gz_C_to_T.fastq Writing a G -> A converted version of the input file cvir_bsseq_all_pe_R2.fastq.gz to cvir_bsseq_all_pe_R2.fastq.gz_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file cvir_bsseq_all_pe_R2.fastq.gz (27591428 sequences in total) Input files are cvir_bsseq_all_pe_R1.fastq.gz_C_to_T.fastq and cvir_bsseq_all_pe_R1.fastq.gz_G_to_A.fastq and cvir_bsseq_all_pe_R2.fastq.gz_C_to_T.fastq and cvir_bsseq_all_pe_R2.fastq.gz_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/sam/data/C_virginica/genomes/GCF_002022765.2_C_virginica-3.0/ with the specified options: -q --score-min L,0,-0.6 -p 28 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 Now starting a Bowtie 2 paired-end alignment for CTread1GAread2CTgenome (reading in sequences from cvir_bsseq_all_pe_R1.fastq.gz_C_to_T.fastq and cvir_bsseq_all_pe_R2.fastq.gz_G_to_A.fastq, with the options: -q --score-min L,0,-0.6 -p 28 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: HWI-C00124:321:CC781ANXX:2:1101:1232:2155_1:N:0:GATCAG/1 77 * 0 0 * * 0 0 ATTGTATTAATTATAAAAAATATNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNATTTATTATTAAAAATATTAAATA FFFFFFFFFBFFFFFFFFFFFFF#################################<>> Writing bisulfite mapping results to cvir_bsseq_all_pe_R1_bismark_bt2_pe.bam <<< Reading in the sequence files /gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/cvir_bsseq_all_pe_R1.fastq.gz and /gscratch/scrubbed/samwhite/outputs/20190312_cvir_gonad_bismark/cvir_bsseq_all_pe_R2.fastq.gz Processed 1000000 sequence pairs so far Processed 2000000 sequence pairs so far Processed 3000000 sequence pairs so far Processed 4000000 sequence pairs so far Processed 5000000 sequence pairs so far Processed 6000000 sequence pairs so far Processed 7000000 sequence pairs so far Processed 8000000 sequence pairs so far Processed 9000000 sequence pairs so far Processed 10000000 sequence pairs so far Processed 11000000 sequence pairs so far Processed 12000000 sequence pairs so far Processed 13000000 sequence pairs so far Processed 14000000 sequence pairs so far Processed 15000000 sequence pairs so far Processed 16000000 sequence pairs so far Processed 17000000 sequence pairs so far Processed 18000000 sequence pairs so far Processed 19000000 sequence pairs so far Processed 20000000 sequence pairs so far Processed 21000000 sequence pairs so far Processed 22000000 sequence pairs so far Processed 23000000 sequence pairs so far Processed 24000000 sequence pairs so far Processed 25000000 sequence pairs so far Processed 26000000 sequence pairs so far Processed 27000000 sequence pairs so far 27591427 reads; of these: 27591427 (100.00%) were paired; of these: 23340236 (84.59%) aligned concordantly 0 times 1930736 (7.00%) aligned concordantly exactly 1 time 2320455 (8.41%) aligned concordantly >1 times 15.41% overall alignment rate 27591427 reads; of these: 27591427 (100.00%) were paired; of these: 23233140 (84.20%) aligned concordantly 0 times 1974165 (7.15%) aligned concordantly exactly 1 time 2384122 (8.64%) aligned concordantly >1 times 15.80% overall alignment rate 27591427 reads; of these: 27591427 (100.00%) were paired; of these: 23225321 (84.18%) aligned concordantly 0 times 1984099 (7.19%) aligned concordantly exactly 1 time 2382007 (8.63%) aligned concordantly >1 times 15.82% overall alignment rate 27591427 reads; of these: 27591427 (100.00%) were paired; of these: 23345816 (84.61%) aligned concordantly 0 times 1923390 (6.97%) aligned concordantly exactly 1 time 2322221 (8.42%) aligned concordantly >1 times 15.39% overall alignment rate Processed 27591427 sequences in total Successfully deleted the temporary files cvir_bsseq_all_pe_R1.fastq.gz_C_to_T.fastq, cvir_bsseq_all_pe_R1.fastq.gz_G_to_A.fastq, cvir_bsseq_all_pe_R2.fastq.gz_C_to_T.fastq and cvir_bsseq_all_pe_R2.fastq.gz_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 27591427 Final Cytosine Methylation Report ================================= Total number of C's analysed: 221887219 Total methylated C's in CpG context: 23206177 Total methylated C's in CHG context: 1030809 Total methylated C's in CHH context: 3473660 Total methylated C's in Unknown context: 127986 Total unmethylated C's in CpG context: 8314955 Total unmethylated C's in CHG context: 49507573 Total unmethylated C's in CHH context: 136354045 Total unmethylated C's in Unknown context: 700229 C methylated in CpG context: 73.6% C methylated in CHG context: 2.0% C methylated in CHH context: 2.5% C methylated in unknown context (CN or CHN): 15.5% Bismark completed in 0d 6h 16m 58s ==================== Bismark run complete ====================