Question: Using unaligned reads as input for trinity
0
gravatar for Pranavathiyani G
17 months ago by
Pondicherry, India
Pranavathiyani G310 wrote:

'I have done HiSAT mapping for my RNAseq sample and I have got unaligned FASTQ sanger file. Can i use this unaligned files as input for running trinity or Should i use the original SRA reads file.

trinity rna-seq assembly • 424 views
ADD COMMENTlink written 17 months ago by Pranavathiyani G310
2

What you should do is to take some of those unaligned reads and check them using blast. At a minimum there may be contamination in data you are using. If so there would be no point in wasting further time on this dataset.

If the reads are from the right genome then you should spend some time figuring out why HISAT2 is unable to align them.

ADD REPLYlink modified 17 months ago • written 17 months ago by genomax89k

Thank you. I'll check that.

ADD REPLYlink written 17 months ago by Pranavathiyani G310
1

In addition to what genomax proposes, I would also just support your approach, or better simply running a full Trinity assembly. It doesn't cost you much except some CPU time and after that you can simply blast all the assembled transcripts. More generally speaking, whenever problems in an analysis arise, one should try out a bunch of different things and make some hypothesis about what is going wrong. Btw, what is your definition of a low mapping rate?

  • wrong genome
  • draft genome with lots of gaps, unmapped regions
  • contamination
  • ribosomal RNA
  • failed sequencing or protocol
ADD REPLYlink modified 17 months ago • written 17 months ago by Michael Dondrup47k

Well technically you can anything you wish. Feed the software with something and it will spit out something more but why do want to do that ?

Share your objective here.

ADD REPLYlink written 17 months ago by lakhujanivijay5.2k

Because the result of HISAT mapping was low.

ADD REPLYlink written 17 months ago by Pranavathiyani G310
1

You should always mention that in your post along with mapping statistics. Anyways, look at this old post, there is a long discussion on similar problem I had faced couple of months back!

Too low mapping percentage using HISAT2 on human reference genome.

ADD REPLYlink written 17 months ago by lakhujanivijay5.2k

Thanks. I'll have a look at it.

ADD REPLYlink written 17 months ago by Pranavathiyani G310
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