Question: Too many intronic variants in RNASeq, is this ok?
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gravatar for Sharon
2.9 years ago by
Sharon490
Sharon490 wrote:

Hi every one

I did GATK somatic mutation pipeline, then I annotated the resulted mutect2 vcf with VEP. 94.32% of mutect2.vcf variants are annotated as intergenic and intronic, and the rest is exonic or ncRNA exonic, is this normal with rnaseq variant calling and somatics? I have many reads discarded by mutect2 though.

Thanks

rna-seq somatic mutations • 793 views
ADD COMMENTlink written 2.9 years ago by Sharon490

Look more closely at your data. Depending on the alignment methods, maybe you have bits of RNA-transcript hanging off the end of an exon, and your tools see intronic mutations. I have seen piles of artifact intronic variants when the read was not trimmed from the end of an exon. It's not really there, but a problem with alignment.

ADD REPLYlink written 2.9 years ago by karl.stamm3.9k

Thanks karl so much. I will check the alignments with IGV.

ADD REPLYlink modified 2.9 years ago • written 2.9 years ago by Sharon490

Are you following guidelines for this? I don't think gatk somatic is appropriate for RNA-seq but might be wrong...

ADD REPLYlink written 2.9 years ago by WouterDeCoster45k

I am using GATK and mutect2, why it is not appropriate?

ADD REPLYlink written 2.9 years ago by Sharon490

I believe those are optimised for DNA sequencing such as exome sequencing.

ADD REPLYlink written 2.9 years ago by WouterDeCoster45k

Sharon, what did you conclude from your data? I am having a similar problem with my rnaseq variant calling analysis (aprox 70% intergenic or intronic).

ADD REPLYlink modified 2.3 years ago • written 2.3 years ago by pereyra.s0
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