Hi,

it seems that your RNA is pretty much degenerated. Either the samples were stored improperly or you have RNAse contamination in your setup. Start with a fresh test sample and fresh kits all over again.

Best

How much RNA are you expecting to get out of plasma? It's essentially a cell-free biofluid, so not exactly transcriptionally active. Your sources of RNA will be either free RNA released from damaged cells, microRNAs, and probably stuff in exosomes/microvesicles.

There won't be a lot.

This paper looks into this in a bit more detail,

Article Human plasma and serum extracellular small RNA reference pro...

And reckons miRs are ~80% of total plasma RNA, with a total conc of ~5-10pM.

Assuming a length of 21nt, that works out to around 30-60pg.ml-1, so you're really not looking at a lot, even if all the other 20% is much longer RNAs.

I don't think the nanodrop is actually telling you anything useful here: I suspect a sample of just guanidium salt contaminants (which absorb at 230) would produce a similar curve.

You have two options:

One, use something with greater sensitivity, like ribogreen/Qbit

Two, just push ahead with your "maybe RNA" and see if you get anything useful downstream.

I agree with John Hildyard that you are probably just looking at contaminants. Even if your RNA was degraded to nucleotides you would still get a 260 absorbance peak, but there is no peak present. I think you just have some absorbing contaminant that is neither protein nor nucleic acid.

Thank you everyone

Hi

The yield obtained depends on technique used after the whole blood sample was gotten from -80. The sample should be thawed quickly on Aluminium blocks at room temperature and further processed using the normal technique your lab adopted. Avoid usage of water bath as it takes long time and leads to more RNA Degradation, this improves RNA quality and yield .