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

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

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

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