Question: HISAT2 tmo and rna-strandness options not working as expected
1
gravatar for DVA
11 months ago by
DVA500
United States
DVA500 wrote:

I am new to HISAT2 and trying to play around with its options. I found the following issues and would appreciate your advice:

  1. I downloaded the genome_tran (for grch38) folder, and wanted to check out the option: --tmo. According to the manual, it should "Report only those alignments within known transcripts.". However, this option allows <10% of my reads to align, while not using this option more than 80% of the reads could get aligned.

    So I went further to examine the reads that could not get aligned after using --tmo, as far as I can manually tell, many of them are from coding regions. What do I miss here - any suggestions?

  2. I also applied --rna-strandness F and --rna-strandness R respectively (with --tmo option on). My reads should be F. However, F and R gave me exactly the same output, with only diffice at " XS:A:" - one is +, the other gives - for a same read.

Thank you very much

rna-seq • 607 views
ADD COMMENTlink modified 11 months ago • written 11 months ago by DVA500
4
gravatar for h.mon
11 months ago by
h.mon24k
Brazil
h.mon24k wrote:

About --tmo, the issue has been asked and reported before, but it seems unresolved. It seems it works better with HISAT2 2.0.4.

About the --rna-strandness, I think this is the described and intended output:

With this option being used, every read alignment will have an XS attribute tag: '+' means a read belongs to a transcript on '+' strand of genome. '-' means a read belongs to a transcript on '-' strand of genome.

Probably this information is used for downstream analyses, such as Stringtie and / or Ballgown.

ADD COMMENTlink modified 11 months ago • written 11 months ago by h.mon24k
2

Wooo thank you! This is of great help!

ADD REPLYlink written 11 months ago by DVA500
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