annealing oligos

Dr. Duncan Clark Duncan at nospam.demon.co.uk
Thu Jan 27 04:10:18 EST 2000


In article <86nnan$tmo$1 at nnrp1.deja.com>, jen_the_glick at my-deja.com
writes
>So when I heat-inactivate my restriction enzymes, should I be cooling
>very slowly, as I did before?  Otherwise, might I be undoing the good I
>did in the initial annealing reaction?

Can you see what the annealing temp would be of the cut down oligos
using say a primer design program? Assuming to left 6 bp before the RE
sites at either end then and providing it isn't AT rich DNA you should
be OK. Don't forget that 65C for 20 mins may not be enough to inactivate
a lot of RE's. An alternative to this may be to use StrataClean Resin
from Stratagene (if they still sell it and if it doesn't latch onto
small DNA). 

Whenever we have to do this we design the oligos such that when
annealed, the appropriate RE overhangs are automatically created. Much
easier! 

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

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




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