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I am isolation RNA from Drosophila larvae brain with TRIzol method. What percentage of extracted RNA will be ribosomal RNA? I am only interested in mRNA, so I am trying to figure out whether I need to get rid of rRNA. Thanks

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  • $\begingroup$ Percentage of what? Total cell mass/volume? Of total RNA moieties? Of total nucleic acids? $\endgroup$ – terdon Feb 25 '14 at 12:53
  • $\begingroup$ Ratio of ribosomal RNA in total RNA if I rephrase it. $\endgroup$ – golgicik Feb 25 '14 at 12:57
  • $\begingroup$ OK, please edit your question to something like "I am extracting RNA using methodX, what percentage of extracted RNA will be ribosomal?" The percentages will also depend on the method used to extract the RNA. $\endgroup$ – terdon Feb 25 '14 at 13:03
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The protocol you are using will not only leave the sample with rRNA but also non coding RNA.

Many RNA protocols will separate mRNA by affinity of a carrier to the polyA tail. This protocol references an older paper that estimates that only 5% of RNA is mRNA. I'd be surprised if this ratio changed by more than 2-3 fold in drosophila.

I assume that %age is by weight but it could be a densitometry measurement which is similarly interpreted.

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  • $\begingroup$ Thanks! As a subsequent question: Do you think that huge amount of rRNA will affect my downstream experiments such as RT-qPCR? I am only interested in mRNAs. $\endgroup$ – golgicik Feb 26 '14 at 9:39
  • $\begingroup$ I've not done qPCR. I would not hesitate to compare multiple standard protocols to get a better sample. Looking at a couple of qPCR protocols you're right I only see Triazol preparations used. PCR is pretty efficient as long as the primers are specific. Some more homework before doing the experiment won't hurt you :) $\endgroup$ – shigeta Feb 26 '14 at 14:35

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