Colony PCR

Namjin Chung nc1 at acpub.duke.edu
Mon Oct 7 19:11:58 EST 1996


On Thu, 3 Oct 1996, Duncan Greig wrote:

> I am trying to PCR up a 2kb fragment, and am having difficulty getting it
> to appear using colony PCR I have had good results with the primers after
> making DNA, but it would be convinient to be able to use colonies.
> 
> Does anyone have a protocol that has produced such large fragments reliably?
> 
> thanks.
> 
> Aidan Weatherill.
> 

Dear Greig:

There are two ways: 1) boil colony which should be picked with pipet tips 
in 1XPCR buffers for 5min; 2)incubate colonies in small amount of 
zymolyase solution for 5min.  After the either way of breaking yeast 
cells, you could do PCR as usual.  It works fine.  By the way, I have 
used Pfu polymerase and its buffer (no affiliation to Stratagene), and I 
just haven't tested with other polymerases.  Theoretically there should 
be no problem with others.  Cheers!


************************************************************************
* Namjin Chung                             *                           *
* Program in Molecular Cancer Biology      * Email: nc1 at acpub.duke.edu *
* Duke University Medical Center, Box 3345 * Voice: +1 (919) 684-2363  *
* Durham, NC 27710                         * Fax  : +1 (919) 681-8253  *
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