hroychow at NMSU.EDU
Mon Jun 8 16:39:55 EST 1998
At 04:39 PM 6/8/98 GMT, jalali at my-dejanews.com wrote:
>I need to blunt end a linearized plasmid with a 5'
>overhang of 2 bases, as well as a fragment of DNA with a 5' overhang.
>The strategy I'm considering at the moment is digestion with Mung-bean
>nuclease. Any thoughts?
>Also, assuming this strategy is okay, will I need to kinase, so that I can
>Any info would be greatly appreciated.
>Mushroom Research Group
>University of Toronto at Mississauga
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No No! Use Klenow and dNTP's instead. The MB nuclease is tricky and you may
end up chewing into the piece. The protocol is simple and quick (15-20 min)
and you can find it in Maniatis or the Red Book. For 3' overhangs, use T4
No need to kinase the plasmid after polishing.
Dr. Hiranya Sankar Roychowdhury
Plant Genetic Engineering Lab.
New Mexico State University
Las Cruces, NM 88003
Ph. (505) 646-5785
hroychow at nmsu.edu
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