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-   -   Bioanalyzer RNA Nano with total RNA - High MW peak above 28S? (http://seqanswers.com/forums/showthread.php?t=72860)

dlv04c 12-02-2016 06:42 AM

Bioanalyzer RNA Nano with total RNA - High MW peak above 28S?
 
I'm getting a significant high MW peak on a nano RNA chip with total RNA. Any thoughts on what this might be caused by? It is present in all the total RNA samples I loaded on this chip.


http://www.bio.fsu.edu/~dvera/highMw...ioanalyzer.png

Amar 12-06-2016 11:39 PM

Possibly genomic DNA. Have you digested your DNA and if so with what?

Some good info here:
http://www2.jabsom.hawaii.edu/genomi...rpretation.pdf

dlv04c 12-07-2016 10:40 AM

it was not denatured
 
Turns out the RNA was not denatured prior to loading on the chip.

marta.m_ 10-24-2017 07:38 AM

1 Attachment(s)
Did it help? You don't see this peak anymore?
I also have problem with this peak (though it is a bit smaller in my case). I am always denaturing my RNA prior to the run but maybe 2 min is not enough?

I think it is not DNA as second DNase treatment did not help and qPCR (+RT vs -RT, housekeeping gene) shows that the RNA is free of DNA (no amplification after 40 cycles). I also tried spiking in a small amount of gDNA and it never resulted in such electropherogram.


Then again I have seen this peak in many publications and examples of "good RNA".
http://www.nature.com/app_notes/nmet...eth908_F1.html
http://genomics.nchresearch.org/submitting_RNA.html
http://www.sigmaaldrich.com/technica...ssessment.html fig 7.9
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4535892/ fig 1.

What can that be?
I am very puzzled.


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