on the left there is the reference profile from Agilent - on the right there is my sample - I don't know if it's realistic to see a clear miRNA-fraction on the Bioanalyzer.
Your RNA quality seems really good, so: any biological reason for such low miRNA fraction? It is known that miRNA acts differently to mRNA in terms of concentrations distribution but...wow: your quantities are so low... If you'll ever decide to construct the library I'll be very interested in known the results (qualitatively). You will check the library length distibution with a DNA chip before the seq run anyway, don't you?
Thank you for the really nice question!
Dear Stefano,
thanks for your reply. It's RNA from another lab, I don't know about the isolation-technique... Yes I will definately run a DNA-chip before sequencing. Today I will screen other RNA-samples in comparison first...
It is possible that the RNA in the reference profile from Agilent has been prepared to be enriched for small RNAs. I have done exosome and RNA co-IP preps where the miRNA peak is almost non-existent on the electropherogram, but I have been able to do microarrays and NGS on them anyway. In these cases I find it more productive to look at the gel-like image and put it on to enhanced gel view, and in this way you will be able to visualise even very subtle bands in this size range.
Thanks for that advice, Belinda - the enhanced gel view did not show much either. I will prepare librarys with different samples now, including one of the critical ones in comparison, maybe it will work as well - like in your case, I will let you know!
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