Question: how to extract intron coordinates (in bed) from bam
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gravatar for praasu
6 months ago by
praasu30
Prague, Czech republic
praasu30 wrote:

I have aligned raw RNA-seq reads against a reference genome with STAR. I would like to extract the intronic coordinates from obtained bam files. Any suggestion?

rna-seq • 376 views
ADD COMMENTlink modified 6 months ago by Devon Ryan96k • written 6 months ago by praasu30
1
gravatar for Devon Ryan
6 months ago by
Devon Ryan96k
Freiburg, Germany
Devon Ryan96k wrote:

This isn't information that one extracts, since it's not held within the BAM files. Instead your steps will be:

  1. Determine where transcripts and exons are (e.g. with stringTie).
  2. Process the resulting GTF file to retrieve introns (using whatever definition of this you'd like).

The second step may require some custom code, depending on whether you want introns that overlap exons (e.g., due to having multiple transcripts for a gene) or not.

ADD COMMENTlink written 6 months ago by Devon Ryan96k

Hi Ryan, The GTF file, that I obtained from the stringTie, is missing strand information. I need strand information to check the splice sites sequence (both at 5' and 3')
Do you have idea, how can I deal with that ?

ADD REPLYlink modified 4 weeks ago • written 4 weeks ago by praasu30

Did you use a strand-specific library prep?

ADD REPLYlink written 4 weeks ago by Devon Ryan96k

We didn't use strand-specific library prep.

ADD REPLYlink written 4 weeks ago by praasu30
1

Then you can't know which strand the transcripts are from.

ADD REPLYlink written 4 weeks ago by Devon Ryan96k

Hi Ryan,

Today, I got to know that mRNA library is stranded. In that case, how can I obtained the strand information or why could it be missing ?

Prasoon

ADD REPLYlink written 28 days ago by praasu30
1

You need to tell your aligner and stringTie that.

ADD REPLYlink written 28 days ago by Devon Ryan96k
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