synthetic RNA stability/degradation

robynb robynb at westmed.wh.su.edu.au
Wed Dec 14 18:22:25 EST 1994


I have a problem with synthetic RNA seemingly breaking down upon
storage.

I have been using any techniques I know of to keep the RNA free from
RNases, such as nuclease-free water (Promega), filter pipette tips,
nuclease-free tubes -which I do not treat in any other way-, addition
of guanidium thiocyanate to the end stage of transcription and rRNasin
is added to the RT mix.

However, when I store the RNA in working aliquots, at -70oC, the first
couple of times I do an RT-PCR (using a fresh aliquot each time) it
seems to work fine, but in subsequent RT-PCRs I seem to lose the PCR
product.

Does anyone have a clue about what could be happening? I would
appreciate any suggestions at all. Please contact me through my email
address or via this news group.

Robyn Biti
Dept. Immunology, Westmead Hospital. Australia.
ROBYNB at westmed.wh.su.edu.au



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