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Check is RNA-seq mapping (tophat) is performed ok (hg38)?

Hi,

I have used default options in Galaxy (just set the Mean Inner Distance between Mate Pairs to 50 which I did not sure it is ok) to map my RNA-seq library to Human genome hg38. Is there a way to check is the mapping good and ok, for example check a housekeeping gene or a gene what I can see how should look like in IGV?

tophat rna-seq
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updated by Ram Ram • written by manekineko manekineko

just open your bam file in IGV and check several genes. Don't forget to index it before open it in IGV (samtools index alignment_file.bam)

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written by Nicolas Rosewick Nicolas Rosewick

Seems OK to me. Are your data stranded?

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updated by Ram Ram • written by Nicolas Rosewick Nicolas Rosewick

R1 read is forwarded and R2 is reverse.

I mainly was worried for setting mean-inner-mate option in tophat as I have problems calculating it. I just know that my reads are 101bp, and the RNA was selected from fragments 300-400.So do not know how much a wrong -r option will harm the mapping. I see from the mapping there is a lot reads overlapping mates

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updated by Ram Ram • written by manekineko manekineko

1 answer

For a comprehensive check, I recommend using Picard's CollectRnaSeqMetrics tool.

IGV, like NicoBxl suggested, is a great way to check individual genes. Galaxy also has its own visualization solution called Trackster.

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written by Dan D Dan D

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