Low copy number vector

Dr. Duncan Clark Duncan at nospam.demon.co.uk
Thu Apr 29 03:51:44 EST 1999


In article <7g7jq7$a28$2 at Masala.CC.UH.EDU>, Michael Benedik
<benedik at uh.edu> writes
>You are getting too fancy on us Duncan with your operator deletion
>trick. Have you tried adding glucose to get catabolite repression in
>BL21?

It is a neat trick and I've been using it for three years or so without
fail. It's nice because you can get blue/white selection without having
to switch on your gene with potentially harmful effects. The other way
is to tag your sequence with lacO and clone that, selecting for blues.

I'm making an HB101(DE3) lysogen today so that may sort it, however I
have slight worries about accidently titrating out all the lacI so
switching on the T7 RNA pol on the DE3 lysogen (which also has lacI) but
that's another day. I think the safest one will be a blue white lacO
arabinose promoter vector, which in theory I should have if the ligated
construct I electroporated last night is OK.

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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