Bowtie2 paired end alignments do not match

[Azufre451]

Hi all,

Maybe someone already encoutered this issue... I am trying to use bowtie2 to map some paired-end reads against a set of assembled contigs (the contigs were built using those reads with SPAdes).

I am using bowtie2 in this way:

bowtie2 --threads 10 --end-to-end --no-unal -x contigs -1  R1.fastq -2 R2.fastq | samtools view -bS - > bams_bowtie2.3/my.bam

However, when inspecting the SAM/BAM (either with Tablet, IGV or manually) I see many cases in which the two mates do not align on the same contig.

I tried to look at the 3rd and 7th fields of the SAM files (the contig-names to which the two mates of the pair align), but the information are discordant. Let's say that I have an alignment that says:

R1/1 -> contig_1 | R1/2 -> contig_2

then I would expect to find another record where

R1/2 -> contig_2 | R1/1 -> contig_1

However, this isn't always true (and for some of the samples I am mapping, it rarely is!).

I also opened an issue/question on the official bowtie2 repository (here) with more detailed explaination of the problem.

I am pretty sure I am missing something important in how to use Bowtie2 on paired-end data... where is the problem?

Thank you very much for your help!
Moreno