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/ (absolute path is '/gscratch/srlab/sam/data/C_virginica/genomes/)' Processing sequences up to read no. 10000000 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/20190222_cvirginica_pe_bismark/subset_10000000'): /gscratch/scrubbed/samwhite/data/C_virginica/BSseq/cvir_bsseq_all_pe_R1.fastq.gz /gscratch/scrubbed/samwhite/data/C_virginica/BSseq/cvir_bsseq_all_pe_R2.fastq.gz Library is assumed to be strand-specific (directional), alignments to strands complementary to the original top or bottom strands will be ignored (i.e. not performed!) Setting parallelization to single-threaded (default) Current working directory is: /gscratch/scrubbed/samwhite/outputs/20190222_cvirginica_pe_bismark/subset_10000000 Now reading in and storing sequence information of the genome specified in: /gscratch/srlab/sam/data/C_virginica/genomes/ Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are /gscratch/scrubbed/samwhite/data/C_virginica/BSseq/cvir_bsseq_all_pe_R1.fastq.gz and /gscratch/scrubbed/samwhite/data/C_virginica/BSseq/cvir_bsseq_all_pe_R2.fastq.gz Input files are in FastQ format Processing reads up to sequence no. 10000000 from /gscratch/scrubbed/samwhite/data/C_virginica/BSseq/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 Created C -> T converted version of the FastQ file cvir_bsseq_all_pe_R1.fastq.gz (10000001 sequences in total) Processing reads up to sequence no. 10000000 from /gscratch/scrubbed/samwhite/data/C_virginica/BSseq/cvir_bsseq_all_pe_R2.fastq.gz 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 G -> A converted version of the FastQ file cvir_bsseq_all_pe_R2.fastq.gz (10000001 sequences in total) Input files are cvir_bsseq_all_pe_R1.fastq.gz_C_to_T.fastq and cvir_bsseq_all_pe_R2.fastq.gz_G_to_A.fastq (FastQ) Now running 2 instances of Bowtie 2 against the bisulfite genome of /gscratch/srlab/sam/data/C_virginica/genomes/ 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: M03631:341:000000000-BLY9V:1:1101:13318:1785_1:N:0:32/1 99 NC_035780.1_CT_converted 13669642 0 76M = 13669704 138 TTTAGTTTTAGTTTGTATATTTTTTTTTAGTTGTATTTTTTTTTATAATTTTATATTAAGATGTGTGTGTAAGAAT CCCCCGGGGGGGGGGGFFCFGGGGGGGG8,CE,C,CEEFFEFEG,,,,>> Writing bisulfite mapping results to cvir_bsseq_all_pe_R1_bismark_bt2_pe.bam <<< Reading in the sequence files /gscratch/scrubbed/samwhite/data/C_virginica/BSseq/cvir_bsseq_all_pe_R1.fastq.gz and /gscratch/scrubbed/samwhite/data/C_virginica/BSseq/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 10000000 reads; of these: 10000000 (100.00%) were paired; of these: 9252606 (92.53%) aligned concordantly 0 times 338002 (3.38%) aligned concordantly exactly 1 time 409392 (4.09%) aligned concordantly >1 times 7.47% overall alignment rate 10000000 reads; of these: 10000000 (100.00%) were paired; of these: 9252985 (92.53%) aligned concordantly 0 times 337570 (3.38%) aligned concordantly exactly 1 time 409445 (4.09%) aligned concordantly >1 times 7.47% overall alignment rate Processed 10000000 sequence pairs so far Processed 10000000 sequences in total Successfully deleted the temporary files cvir_bsseq_all_pe_R1.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: 10000000 Final Cytosine Methylation Report ================================= Total number of C's analysed: 23154065 Total methylated C's in CpG context: 2018853 Total methylated C's in CHG context: 83895 Total methylated C's in CHH context: 571199 Total methylated C's in Unknown context: 6834 Total unmethylated C's in CpG context: 1311294 Total unmethylated C's in CHG context: 4958974 Total unmethylated C's in CHH context: 14209850 Total unmethylated C's in Unknown context: 78717 C methylated in CpG context: 60.6% C methylated in CHG context: 1.7% C methylated in CHH context: 3.9% C methylated in unknown context (CN or CHN): 8.0% Bismark completed in 0d 0h 31m 42s ==================== Bismark run complete ====================