quantifying ribosomal content in Illumina libraries
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6.9 years ago
Richard ▴ 580

Hi all,

Is there a standard tool people use to quantify the relative amounts of Ribosomal reads in RNA-seq libraries?

 

ribosomal • 1.3k views
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6.9 years ago
Danielk ▴ 620

Yes, you can use picards ColleceRnaseqmetrics. 

https://broadinstitute.github.io/picard/command-line-overview.html#CollectRnaSeqMetrics

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Does everyone use the same set of Ribosomal intervals on hg19, or is there some variation between group?
 

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I use these:

https://github.com/dakl/genome

I've also seen people get them from the ucsc mysql dB. I guess they give very similar results. 

 

 

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Hmmm. This list doesn't seem to include 28S or 18S which I've read are the great majority of detected rRNAs

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You're right. In GRCh37, it maps to an unplaced contig. However, using these regions does distinguish between good and bad ribodepletions. With a good depletion, we get 0-2% rRNA content, and i our failed experiments with a robotic protocol with a bad sometimes see as high as 30%. With TruSeq Stranded Total RNA with Ribo-Zero Gold, we consistently get 0-2%. 

But you are right, these values should be interpreted as relative within a series, rather than estimates of absolute rRNA content. 

 

 

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