Bowtie seems to be working fine (tested command '/Applications/bioinfo/bowtie2-2.3.4.1-macos-x86_64/bowtie2 --version' [2.3.4]) Output format is BAM (default) Alignments will be written out in BAM format. Samtools found here: '/Applications/bioinfo/samtools-1.3.1/samtools' Reference genome folder provided is /Volumes/Serine/wd/18-03-15/genome/ (absolute path is '/Volumes/Serine/wd/18-03-15/genome/)' Processing sequences up to read no. 10000 from the input file Input files to be analysed (in current folder '/Volumes/Serine/wd/18-04-12/tmp'): /Volumes/Serine/wd/18-04-10/zr2096_5_s1_R1.fastq /Volumes/Serine/wd/18-04-10/zr2096_5_s1_R2.fastq 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: /Volumes/Serine/wd/18-04-12/tmp Now reading in and storing sequence information of the genome specified in: /Volumes/Serine/wd/18-03-15/genome/ Single-core mode: setting pid to 1 Paired-end alignments will be performed ======================================= The provided filenames for paired-end alignments are /Volumes/Serine/wd/18-04-10/zr2096_5_s1_R1.fastq and /Volumes/Serine/wd/18-04-10/zr2096_5_s1_R2.fastq Input files are in FastQ format Processing reads up to sequence no. 10000 from /Volumes/Serine/wd/18-04-10/zr2096_5_s1_R1.fastq Writing a C -> T converted version of the input file zr2096_5_s1_R1.fastq to zr2096_5_s1_R1.fastq_C_to_T.fastq Writing a G -> A converted version of the input file zr2096_5_s1_R1.fastq to zr2096_5_s1_R1.fastq_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file zr2096_5_s1_R1.fastq (10001 sequences in total) Processing reads up to sequence no. 10000 from /Volumes/Serine/wd/18-04-10/zr2096_5_s1_R2.fastq Writing a C -> T converted version of the input file zr2096_5_s1_R2.fastq to zr2096_5_s1_R2.fastq_C_to_T.fastq Writing a G -> A converted version of the input file zr2096_5_s1_R2.fastq to zr2096_5_s1_R2.fastq_G_to_A.fastq Created C -> T as well as G -> A converted versions of the FastQ file zr2096_5_s1_R2.fastq (10001 sequences in total) Input files are zr2096_5_s1_R1.fastq_C_to_T.fastq and zr2096_5_s1_R1.fastq_G_to_A.fastq and zr2096_5_s1_R2.fastq_C_to_T.fastq and zr2096_5_s1_R2.fastq_G_to_A.fastq (FastQ) Now running 4 individual instances of Bowtie 2 against the bisulfite genome of /Volumes/Serine/wd/18-03-15/genome/ with the specified options: -q --score-min L,0,-1.2 -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 zr2096_5_s1_R1.fastq_C_to_T.fastq and zr2096_5_s1_R2.fastq_G_to_A.fastq, with the options: -q --score-min L,0,-1.2 -p 4 --reorder --ignore-quals --no-mixed --no-discordant --dovetail --maxins 500 --norc)) Found first alignment: HWI-C00124:321:CC781ANXX:1:1101:1218:2142_1:N:0:CAGATC/1 77 * 0 0 * * 0 0 NAATAAAATATTATTTAAATTTTATTATTTATANNNNNNNNNNNNNNNNNNATTNTTTTTATTATATATATAAAATTATTTTAAAATTATTATATTTTTT #<>> Writing bisulfite mapping results to zr2096_5_s1_R1_bismark_bt2_pe.bam <<< Reading in the sequence files /Volumes/Serine/wd/18-04-10/zr2096_5_s1_R1.fastq and /Volumes/Serine/wd/18-04-10/zr2096_5_s1_R2.fastq 10000 reads; of these: 10000 (100.00%) were paired; of these: 7135 (71.35%) aligned concordantly 0 times 1199 (11.99%) aligned concordantly exactly 1 time 1666 (16.66%) aligned concordantly >1 times 28.65% overall alignment rate 10000 reads; of these: 10000 (100.00%) were paired; of these: 7197 (71.97%) aligned concordantly 0 times 1225 (12.25%) aligned concordantly exactly 1 time 1578 (15.78%) aligned concordantly >1 times 28.03% overall alignment rate 10000 reads; of these: 10000 (100.00%) were paired; of these: 7193 (71.93%) aligned concordantly 0 times 1232 (12.32%) aligned concordantly exactly 1 time 1575 (15.75%) aligned concordantly >1 times 28.07% overall alignment rate 10000 reads; of these: 10000 (100.00%) were paired; of these: 7151 (71.51%) aligned concordantly 0 times 1191 (11.91%) aligned concordantly exactly 1 time 1658 (16.58%) aligned concordantly >1 times 28.49% overall alignment rate Processed 10000 sequences in total Successfully deleted the temporary files zr2096_5_s1_R1.fastq_C_to_T.fastq, zr2096_5_s1_R1.fastq_G_to_A.fastq, zr2096_5_s1_R2.fastq_C_to_T.fastq and zr2096_5_s1_R2.fastq_G_to_A.fastq Final Alignment report ====================== Sequence pairs analysed in total: 10000 Final Cytosine Methylation Report ================================= Total number of C's analysed: 176153 Total methylated C's in CpG context: 15197 Total methylated C's in CHG context: 1267 Total methylated C's in CHH context: 4823 Total methylated C's in Unknown context: 673 Total unmethylated C's in CpG context: 7270 Total unmethylated C's in CHG context: 37436 Total unmethylated C's in CHH context: 110160 Total unmethylated C's in Unknown context: 2631 C methylated in CpG context: 67.6% C methylated in CHG context: 3.3% C methylated in CHH context: 4.2% C methylated in unknown context (CN or CHN): 20.4% Bismark completed in 0d 0h 0m 26s ==================== Bismark run complete ====================