This is a test version of Biostars. For the public version, visit https://www.biostars.org.
diagnosing contamination in RNA-seq data

I have some bulk RNA-seq data on human samples that have clearly been contaminated with some non-human sources, I'm observing dismal alignment to the human genome even after generously adjusting thresholds.

I want to diagnose the source of contamination at a broad level (e.g. where are these reads coming from?)

Originally I was going to BLAST except it takes way too long and is overkill on my question (I'm wondering what non-human sources are there, not the genes).

Are there any basic packages that people know of that offer species/genuses of RNA-seq data that I can bake into my existing QC pipeline?

thanks!

sequencing ngs bulk nonhuman
• 2,910 views • link
updated by igor igor • written by wiscoyogi • wiscoyogi

use fastqscreen. It will screen for contamination for model organisms, human, mouse, rat and vectors by default. If you can guess the contaminant source organism, index the genome, place indices it in appropriate location, edit the config and fastqscreen will screen fastq against those genomes too.

• 0 views • link
written by cpad0112 cpad0112

BLAST is definitely not the best tool for the job. There are a few alternatives covered here: Faster BLAST alternative

• 0 views • link
written by igor igor

BLAST is more sensitive than all these alternatives.

• 0 views • link
written by shelkmike shelkmike

Yes, but you can't realistically BLAST thousands or millions of reads. The goal here is to "diagnose the source of contamination at a broad level".

• 0 views • link
written by igor igor

2 answers

I would suggest you use something based on k-mers. Maybe this article gives an idea.

• 0 views • link
written by supertech supertech

To diagnose contamination I align random 100 or 1000 reads to NCBI nt (downloaded from https://ftp.ncbi.nlm.nih.gov/blast/db/) by Blastn and look at taxons of best matches.

• 0 views • link
written by shelkmike shelkmike

Log in to answer this question.

More posts like this