Is this error forgivable?
hansw at sci.kun.nl
Sun Aug 25 06:27:52 EST 2002
iayork at panix.com (Ian A. York) wrote in message news:<ak7vbg$88b$1 at reader1.panix.com>...
> In article <20020824081117.12380.00003288 at mb-fm.aol.com>,
> JSUPolek <jsupolek at aol.com> wrote:
> >Added 2X the concentration of ampicillin to my LB agar that I was supposed to
> >(100ug/ml v. 50 ug/ml). Using DH5-alpha and pGEM. Any reason not to use them?
> > Let them incubate a little longer? Thanks
> Go ahead and use them. If the bugs are Amp-resistant in the first place,
> under these circumstances, they're almost infinitely Amp resistant. By
> the time you see the colonies, they'll have destroyed all the Amp around
> them anyway.
> 100 ug/ml is the standard Amp concentration in a bunch of labs anyway.
I have used the pGEM T easy vector in combination with E. coli XLI
Blue a couple of times using both amp concentrations you have
mentioned. I have noticed no decline in transformation efficiency
(number of CFU counted on plate) and found no reason not to use the
construct. As ian says; a lot of protocols provided with vectors (i.e.
pGEX & pET) recommend the 100 ug/ml amp concentration. But do not
attempt to incubate too long since this will yield satelite formation
which will make the cloning harder to do.
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