Is there a simple test for RNase and DNase free

Dr. Duncan Clark Duncan at nospam.demon.co.uk
Sat Jan 27 05:21:15 EST 2001


In article <Vshc6.249250$59.62471563 at news3.rdc1.on.home.com>, the
eminent EO at Excite at Home - The Leader in Broadband
http://home.com/faster wrote
>We would like to test a samples of the batch for
>"DNASE and RNASE free" to confirm they are
>molecular biology grade.  Even the "protease free" test.

It depends on how free you mean free which then determines what the
sensitivity is of the assay you must use. 

DNase can be checked by incubating a plasmid digest of small fragments
(i.e. 50-500bp) in a 1x LOW RE reaction buffer with the suspect whatever
for 16 hours at 37C, remembering to do a negative control, as well. Run
on acrylamide gel and see if there is any change.

For RNA you can do something similar with a transcript etc.

For protease, Taq pol is a great one for checking pancreatic DNAse to be
protease free. It really gets chewed up when visualised on a protein
gel. Problem is you need 1ug, 250u, per assay. Not a problem for us, a
manufacturer but I'd guess there must be fluorescent protease substrates
that could be used in place.

For increased sensitivity on RNase and DNase use 32P labelled substrates
and run as above and autoradiograph or better still count released 32P.
Very sensitive.

Duncan
-- 
The problem with being on the cutting edge is that you occasionally get 
sliced from time to time....

Duncan Clark
GeneSys Ltd.
Tel: +44(0)1252376288
FAX: +44(0)8701640382
http://www.dnamp.com
http://www.genesys.demon.co.uk






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