I am trying the BISMARK software for Bisulfite sequence analysis but in the second step when I use
REF_DIR=/home/DNA_METHYLATION/REFERENCE bismark --basename DM1 --output_dir OUTPUT_DM1 --samtools_path /home/samtools-1.8 --path_to_bowtie /home/software/bowtie-master --bowtie1 $REF_DIR -1 DM1QualityTrimmed.A_1.fq.gz,DM1QualityTrimmed.B_1.fq.gz,DM1QualityTrimmed.C_1.fq.gz,DM1QualityTrimmed.D_1.fq.gz,DM1QualityTrimmed.E_1.fq.gz -2 DM1QualityTrimmed.A_2.fq.gz,DM1QualityTrimmed.B_2.fq.gz,DM1QualityTrimmed.C_2.fq.gz,DM1QualityTrimmed.D_2.fq.gz,DM1QualityTrimmed.E_2.fq.gz
I only get output files corresponding to the last pair. A bam file named "
DM1_pe.bam" and the following report in the file "
Bismark report for: DM1QualityTrimmed.E_1.fq.gz and DM1QualityTrimmed.E_2.fq.gz (version: v0.20.0) Bismark was run with Bowtie against the bisulfite genome of /home/DNA_METHYLATION/REFERENCE/ with the specified options: -q -n 1 -k 2 --best --maxins 500 --chunkmbs 512 Option '--directional' specified (default mode): alignments to complementary strands (CTOT, CTOB) were ignored (i.e. not performed) Final Alignment report ====================== Sequence pairs analysed in total: 52869990 Number of paired-end alignments with a unique best hit: 26447546 Mapping efficiency: 50.0% Sequence pairs with no alignments under any condition: 23883829 Sequence pairs did not map uniquely: 2538615 Sequence pairs which were discarded because genomic sequence could not be extracted: 42243 Number of sequence pairs with unique best (first) alignment came from the bowtie output: CT/GA/CT: 13260580 ((converted) top strand) GA/CT/CT: 0 (complementary to (converted) top strand) GA/CT/GA: 0 (complementary to (converted) bottom strand) CT/GA/GA: 13186966 ((converted) bottom strand) Number of alignments to (merely theoretical) complementary strands being rejected in total: 0 Final Cytosine Methylation Report ================================= Total number of C's analysed: 1410585296 Total methylated C's in CpG context: 58687841 Total methylated C's in CHG context: 590155 Total methylated C's in CHH context: 1661886 Total unmethylated C's in CpG context: 16608516 Total unmethylated C's in CHG context: 327902201 Total unmethylated C's in CHH context: 1005134697 C methylated in CpG context: 77.9% C methylated in CHG context: 0.2% C methylated in CHH context: 0.2% Bismark completed in 0d 16h 52m 0s
Could anyone please tell why I do not get 5 bam files and 5 txt(report) files corresponding to the 5 pairs of input files?
Thanks in advance.