Question: Tophat 1.4 Output (Accepted_Hits.Bam)
0
gravatar for thecuriousbiologist
6.7 years ago by
United States
thecuriousbiologist430 wrote:

I have read the Tophat 1.4 manual, however I do not seem to understand the outputs very well.

I used a reference genome for my alignment with Tophat and I got a file called "accepted_hits.bam"

This file's cigar strings don't contain any soft clips (S). Can someone explain me why this is happening ?

Also, this was for the reference genome. If I wanted to align to a junction database, should I just use the junction reference instead of the genome reference ? Or does Tophat align to junctions automatically ?

Very confused with the documentation.

tophat bam output • 3.0k views
ADD COMMENTlink modified 4.8 years ago by Biostar ♦♦ 20 • written 6.7 years ago by thecuriousbiologist430
1
gravatar for Ido Tamir
6.7 years ago by
Ido Tamir5.0k
Austria
Ido Tamir5.0k wrote:
  • tophat splits the reads, but still requires everything to match. You could have mismatches in the beginning or end, but these would be scored as M in the CIGAR string. You have to look in the MD tag for mismatches. Old tophat versions dont have the optional MD tag. You can generate it with samtools calmd.

  • You always need a reference sequence. You can use a junction file or a GTF file in addition to this. Tophat automatically tries to align reads that can not be mapped completely by aligning them in a split fashion, thus discovering junctions. You can turn this off, modify it with parameters or supply your own junctions. But you should also have a junctions.bed file in your output and see some reads with a CIGAR like 20M500N50M.

ADD COMMENTlink modified 6.7 years ago • written 6.7 years ago by Ido Tamir5.0k

Can the 500N part of the CIGAR contain soft clips ?

ADD REPLYlink written 6.7 years ago by thecuriousbiologist430

500N is a gap tophat is marking an intron

ADD REPLYlink written 5.2 years ago by karl.stamm3.5k
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