Hi,
My RNA-seq data has transcript fusions and novel splice-junctions. So, I am trying to use segemehl in local alignment (-S) mode to map it. I am using the -Z 20 (--minfraglen <n> min length of a spliced fragment). However, I get fragments of length <20 nt in my output. The command for mapping looks like this-
~/bin/segemehl/segemehl.x --silent -d reference.fa -q reads.fastq -i reference.idx -o file.sam -S -A 95 -Z 20
Example output,
M02096:114:000000000-AEH6V:1:1106:27332:13940 1:N:0:1 16 XIST 78 255 **9M** * 0 0 **TTTCTCTTG** IIIFIIIII NM:i:0 MD:Z:9 NH:i:1 XI:i:0 XL:i:2 XA:Z:Q XX:i:223 XY:i:231 XQ:i:1 XP:Z:XIST XU:i:990 XS:i:0
Can someone please explain if I am doing something wrong? Why do I see alignments of <20 nt even after specifying -Z 20?
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mihir.metkar •
Dear mihir.metkar,
I talked to one of the developer and he explained it like this:
Even though a min fragment length was specified in some cases the algorithm is shifting smaller parts of a read towards already known junctions! It is a feature!
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written
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David Langenberger
Hi David,
Thank you for the answer. Is there a way to turn the feature off?
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written
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mihir.metkar •
You can postfilter the sam file with Reformat from the BBMap package to get rid of the short fragments:
reformat.sh in=mapped.sam out=filtered.sam minlen=20
It can't be turned off. You can directly contact the developers to change it, or follow Brians idea.
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written
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David Langenberger
Ok, thank you all for your answers.
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written
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mihir.metkar •
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