nuclear vs. total RNA isolation question

Martin Hughes mhughes at
Tue Jul 18 12:31:43 EST 1995

In article <1995Jul17.105850.15117 at wvnvms>, lsalati at says:
>I have a question about determining mRNA abundance in nuclear and
>cytoplasmic fractions.  Note that I'm specifically interested in
>mature message so intron probes would not be useful.

In the 1980s I worked in a lab analysing splicing of polyomavirus RNAs
we routinely seperated nuclei and cytoplasm from tissue culture cells
by NP40 lysate with Macaloid in the extract to inhibit RNase, then spin
thro sucrose cushion followed by proteinase K digestion to kill RNase.
See publications by Robert Kamen around 1980-1982 (ICRF London).
Also, could you use a splice junction probe to detect mature RNA and
how about tRNA or rRNA as controls.     Alison Cowie.

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