TA cloning and competent cells

Dr. Duncan Clark duncan at genesys.demon.co.uk
Wed Jan 15 04:51:27 EST 1997


In article <Pine.A41.3.95.970113183839.20104H-100000 at aix1.uottawa.ca>,
sunfish <s535290 at aix1.uottawa.ca> writes
>Hi all,
>
>does anyone ever use homemade competent cells for use with TA cloning ? 
>(we use the pCR2.1 by invitrogen)  Sometimes we run out of the competent
>cells that come with the kit and if I'm in a hurry and can't wait for new
>cells to arrive, it'd be nice to know that the home made stuff is alright.
>If anyone does use their own competent cells, what strain do you use, and
>what method do you use for making the cells competent ?
>
>thanks in advance,
>
>Ed
>

We use TOP10 (from Invitrogen) or you can use the Invalpha they provide
(same genotype as DH5alpha). You will need an efficiency of above 5 x
10E7 preferably >10E8. Inoue's protocol works fine.

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

Dr. Duncan Clark
DNAmp Ltd.
TEl/FAX 01252376288
http://www.dnamp.com
http://www.genesys.demon.co.uk



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