Competent cells Was: Re: filter-sterilized MilliQ water for PCR?

Richard P Grant rgrant at see.sig.for.address
Fri Sep 19 03:41:39 EST 1997


(I also reponded by email)

The problem with autoclaving competent cell reagents is the danger of
introducing ccDNA, which would cause obvious problems.

Low transformation efficiencies may be due to any of a number of factors,
including the method (!), slow freezing of cells, the cell line, storage,
but probably not autoclaving per se.

I use a modification of Cohen 1972, you might want to check out

http://worf.molbiol.ox.ac.uk/~rgrant/thesis/2materialsandmethods.html

and search for 'competent'.

Richard

Wilson (netson at uxmail.ust.hk) wrote:
} Dear Mr. Grant,
} 	Is it possilbe that contaminants can be introduced into the
} reagents during autoclaving?  I always sterile the reagents (e.g LB,
} glycerol and transformation buffer) for competent cell preps by autoclave
} and the transformation efficiency of my competent cell is often low and
} unstable.  I would like to know the problem whether casued by the
} autoclaving the reagent.  



--
Richard P. Grant  MA  DPhil          University of Oxford | rgrant at molbiol
http://www.molbiol.ox.ac.uk/~rgrant  FFPGP                |    ox ac uk
------# It's highly organized beach sand. Why should it stress me? #------



More information about the Methods mailing list