IUBio

Help about methylation and SexAI

Dr. Duncan Clark Duncan at nospam.demon.co.uk
Fri Mar 12 04:28:47 EST 1999


In article <36E7CC06.1339BA2E at imiucca.csi.unimi.it>, Gianluca Molla
<m at imiucca.csi.unimi.it> writes
>Now I think I have
>to purify my DNA from a E.coli strain that is dcm-, but
>almost every cloning strain in cdm+ (except JM110).
>Can I use this strain or its genotype is not good for cloning
>and plasmid amplification? And what about other methylases
>like EcoRII? Are they a problem?

Get hold of an NEB catalogue at look in the appendix at the back. They
have a page on the effects of dam and dcm methylation on restriction
enzymes whose recognition sequence overlaps these methylation sites.

All standard E.coli K12 strains are both dam+ (methylates A in GATC) and
dcm+ (methylates internal C in CC(A/T)GG). Exceptiosn are JM110, GM48
and some other GM strains. I think you can get these free from NEB with
your order. Again info. Is in the back of the NEB catalogue.

All E.coli B strains are dam+ dcm- so you could use one of those.

EcoR II methylase is the methylase of the plasmid carried EcoR II
restriction modification system and is not present in laboratory
strains.

So use a dcm- E.coli to prepare your plasmid in and it will cut 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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