Kinetic of RNA degradation

Dr. Peter Gegenheimer PGegen at UKans.nolospamare.edu
Fri Aug 4 17:56:57 EST 2000


On Thu, 3 Aug 2000 15:30:46, mlsulliv at facstaff.wisc.edu ("Michael L.
Sullivan") wrote:

ð I was at a biochemistry seminar some time ago where the presenter (who is
ð an RNA biochemist) said that the half life of RNA at pH 7 (I think at pH 7)
ð and room temperature was 7 years!  This seems really stable, but he pointed
ð out that the half life of DNA was 7000 years.  Sorry, I don't have a
ð reference for this.
ð Mike
ð
ð >Can somebody help me : how fast is the degradation of a RNA (say from
ð >20-mer to 100-mer) versus temperature in a buffer that does not contain
ð >RNAse (but sometimes magnesium ions).
ð >
ð >Thanks for your help
ð >
ð >Carmelo

RNA is degraded by OH(-) ions present in water as well as generated from
magnesium hydrate [Mg(2+).6H2O]. The kinetics of hydrolysis are
temperature-dependent. RNA fragment "ladders" are prepared by heating to
90-100C for a few min at pH 8.0 to 10.0 in buffer lacking Mg(2+). However,
this normally gives fragments >= 50nt (as I recall). The size distribution
depends on the length of time.

If you are trying NOT to degrade RNA, keep it at pH 6-7 with 0.1 - 1.0 mM
EDTA. If you want controlled fragmentation, such as a 1-nt-spaced size ladder,
heat 5'-(32)P-RNA in bicarbonate buffer (pH 10) and take aliquots from 1 min
to 10-20 min. I don't have a precise protocol handy. There are some good new
"RNA Protocol" books published within the last 5 years, and Ambion's web site
has lots of RNA protocols.

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