I am mapping mate-pair reads with bowtie2 (I use bwa for most of the applications, but I wasn't happy with the result). I am using following commands
bowtie2 -X 500 -S MPout.sam --fr --very-sensitive -x reference.fasta -1 MPR1.fastq MPR2.fastq
bowtie2 -X 10000 -S MPout.sam --fr --very-sensitive -x reference.fasta -1 MPR1.fastq MPR2.fastq
(I reverse complemented the reads prior to the mapping so fr orientation should be correct)
The results for both commands and 10,000 reads are identical:
2500 reads; of these:
2500 (100.00%) were unpaired; of these:
2168 (86.72%) aligned 0 times
60 (2.40%) aligned exactly 1 time
272 (10.88%) aligned >1 times
13.28% overall alignment rate
This doesn't seem right, correct -X parameter should in my opinion result in higher proportion of reads mapped. Or am I missing any important parameter? I might be, I have never used bowtie2 with mate pair-reads. There shouldn't be any paired-end read contamination in my library (I filtered those out already).
Thanks for help. If you recommend something other than bowtie2, could you please provide command you use for mate-pair reads?