Best mapped reads from Bowtie2 alignment: AS>XS

Question

[Tash]

Hello all,

I have a sam file that I have mapped using Bowtie2, using the following parameters:

bowtie2 -q -p 8 -N 1 -X 1000

I want to get the uniquely mapped reads, as well as the best alignment in the case where there are multiple alignments.

To get the uniquely mapped reads I have done this, and it has worked well:

grep -E "@|NM:" input.sam | grep -v "XS:" > unique.sam

However, for the best alignment in the case of multimappers, when I want anything where AS > XS I have had no luck.

For example, of the four reads below, I would want to keep all reads except for the 3rd, in which the XS > AS.

A00129:1205:HLKCMDSX3:1:1101:3341:1235  99      chr16   74463701        42      150M    =       74463850        299     TGTTTCCTCTCATTTTTAACCATTTATTTTGGTCCATTTTGGCTGGAAAAATTGACCAGTCCAGGTCAATTTGTGATGCTGAAATACTTTGCATGTAGGGCATTTCTGGAGGATGGTTGATCTTCTAGGGGTCATAAGTCAGATACTGGG  FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFF,FFFFFFFFF:FFF:FFFFFF:FFF:FFFFFFFFFFFFFFFFFFFFFF  AS:i:0  XN:i:0  XM:i:0  XO:i:0  XG:i:0  NM:i:0  MD:Z:150        YS:i:-3 YT:Z:CP
A00129:1205:HLKCMDSX3:1:1101:3341:1235  147     chr16   74463850        42      150M    =       74463701        -299    GAAAAAACAATACAAAACAAACAAAAACAAAACACAACAAGCAAAAACTGTTTTCCCTTTTCCCAGCCAGTATAAAAGCCAGTTAAGAACTTGACTAAGGGTAGAACTTTAGACCAGTTTTGAACTTTTTTTTTATTAGATATTTTCTTT  FFFF:FFFF:FFF,F:FFFFFFFFF:FFFF:FFF,,FF:F:FFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF  AS:i:-3 XN:i:0  XM:i:1  XO:i:0  XG:i:0  NM:i:1  MD:Z:34A115     YS:i:0  YT:Z:CP
A00129:1205:HLKCMDSX3:1:1101:4734:1235  99      chr2    98497124        2       150M    =       98497395        421     TGGCAAGAAAACTGAAAATCATGGAAAATGAGAAACATCCACTTGACGACTTGAAAAATGACGAAATCACTAAAAAACCTAAAAAATGAGAAATGCACACTGAAGGACCTGGAATATGGCGAGAAAACTGAAAATCACGGAAAATGAGAA  FFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFF,FFFFFFFFFFFFFFFFFFFFFFFFF  AS:i:-30        XS:i:-36        XN:i:0  XM:i:6  XO:i:0  XG:i:0  NM:i:6  MD:Z:1T37T38G1G7T37C23  YS:i:-34        YT:Z:CP
A00129:1205:HLKCMDSX3:1:1101:19578:1078 83      chr2    98497405        6       150M    =       98496949        -606    AGACACATCCACTTGACGACTTGAAAAATGACGAAATCACTAAAAAACGTGAAAAATGAGAAATGCACACTGAAGGACCTGGAATATGGCGAGAAAACTGAAAATCACGGAAAATGAGAAATACACACTTTAGGACGTTAAATATGGCNA  FFFFF:FFFFFFFFFF,FF:FFFFFFFFFFF,FF::FFFFFFFF,F:FFFFFFFF,FFFFFF::FFFFFFFFFFFFFFF:FFFF,FFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFF:FFFF:FFFFFFFFFF:FFFFF:FFFFFF#F  AS:i:-16        XS:i:-21        XN:i:0  XM:i:4  XO:i:0  XG:i:0  NM:i:4  MD:Z:3A13T120G9G1       YS:i:-36        YT:Z:CP

I'm at a loss for how to do this in bash, so if someone would be happy to point me in the right direction, it would be really appreciated!

Many thanks.

Answer 1

[Pierre Lindenbaum]

not tested, but it could be:

samtools view -e '[AS] && [XS] && [AS] > [XS]'  in.bam

[Tash]

Wonderful Pierre, it worked perfectly to get the best alignment! I merged this with the uniquely mapping reads to get the file I needed. Thank you very much.

[Pierre Lindenbaum]

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