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. 13795713 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/half_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/half_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. 13795713 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 (13795714 sequences in total) Processing reads up to sequence no. 13795713 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 (13795714 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 13795713 reads; of these: 13795713 (100.00%) were paired; of these: 11039686 (80.02%) aligned concordantly 0 times 1258510 (9.12%) aligned concordantly exactly 1 time 1497517 (10.85%) aligned concordantly >1 times 19.98% overall alignment rate 13795713 reads; of these: 13795713 (100.00%) were paired; of these: 11124252 (80.64%) aligned concordantly 0 times 1220466 (8.85%) aligned concordantly exactly 1 time 1450995 (10.52%) aligned concordantly >1 times 19.36% overall alignment rate 13795713 reads; of these: 13795713 (100.00%) were paired; of these: 11032856 (79.97%) aligned concordantly 0 times 1266345 (9.18%) aligned concordantly exactly 1 time 1496512 (10.85%) aligned concordantly >1 times 20.03% overall alignment rate 13795713 reads; of these: 13795713 (100.00%) were paired; of these: 11120790 (80.61%) aligned concordantly 0 times 1225463 (8.88%) aligned concordantly exactly 1 time 1449460 (10.51%) aligned concordantly >1 times 19.39% overall alignment rate Processed 13795713 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: 13795713 Final Cytosine Methylation Report ================================= Total number of C's analysed: 142342778 Total methylated C's in CpG context: 15059429 Total methylated C's in CHG context: 671225 Total methylated C's in CHH context: 2059702 Total methylated C's in Unknown context: 76501 Total unmethylated C's in CpG context: 5460824 Total unmethylated C's in CHG context: 32006667 Total unmethylated C's in CHH context: 87084931 Total unmethylated C's in Unknown context: 435443 C methylated in CpG context: 73.4% C methylated in CHG context: 2.1% C methylated in CHH context: 2.3% C methylated in unknown context (CN or CHN): 14.9% Bismark completed in 0d 3h 44m 47s ==================== Bismark run complete ====================