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Old 03-03-2015, 07:47 AM   #21
dpryan
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Therein lies the problem, there is no single definition of "uniqueness". There are multiple incompatible definitions. Further, if we relax the --score-min settings enough then by some definitions there will never be any unique alignments. This is why MAPQ is a generally more useful concept and you'd be better served just forgetting about the term "unique" in this context.
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Old 03-03-2015, 07:36 PM   #22
am@i
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Thank you @devon
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Old 03-04-2015, 02:03 AM   #23
JPOeyen
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Thanks for the help, I will try to figure out how to best solve the issue for my experiments.

I found this, which might be of interest to others trying to understand how bowtie2 assigns scores: link. There are also some interesting thoughts on uniqueness discussed in this and an older blog post.
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Old 03-30-2017, 07:01 PM   #24
keo
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Hi all,
This worked for me, but I don't know if it is a general solution. If you set the -k paramenter in Bowtie2 to >=2, you should have at least twice the name of the read in your SAM file. You can use that to remove reads that appear >1 times in the file my_filename.sam. This way you don't have to undertand how Bowtie sets tags and flags.
Quote:
prefix="my_filename"
tail -n +$(expr $(grep "^@" "$prefix.sam" | wc -l | cut -f 1 -d " ") + 1) "$prefix.sam" | sort | cut -f 1 | uniq -cd | cut -d " " -f 8 > "$prefix.toremove"
grep -vwF -f "$prefix.toremove" "$prefix.sam" > "$prefix.unique.sam"
rm "$prefix.toremove"
Comments appreciated.

Last edited by keo; 03-30-2017 at 07:18 PM.
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