A Question About Samtools And Calculating Depth

Question

[Sameet]

I have some RNA-Seq data. I want to know if there is a way to use samtools to calculate strand-specific depth from the accepted_hits.bam file generated by tophat.

Answer 1

[Ashutosh Pandey]

Not sure what exactly you mean. Can you be little more descriptive. So far whatever I understand I think you should use Cufflinks for the next step. You can also make use of SAM flags (Check out - http://picard.sourceforge.net/explain-flags.html.) that tell you which strand, forward or reverse a read belongs to ? You can use this information to split your bam file into two - forward and reverse and calculate coverage.

[Sameet]

I do have the cufflinks output for these. But i only wanted to see if i can visualize the alignment itself.

[Ashutosh Pandey]

Read this post. If you need to visualize it on genome browser, you can create strand specific wiggle files as described in the post.

[Sameet]

Thank you @ashutoshmits. This was very helpful.