PCR directly on whole E coli

Ed Beaty ed_beaty at qms1.life.uiuc.edu
Mon May 23 17:28:00 EST 1994


In article <nhl-230594105448 at 132.206.27.203>, nhl at binkley.cs.mcgill.ca
(Nancy Lemieux) wrote:
> 
> 
> Hi!
> 
> I have screened for plasmids from E. coli colonies.  I take a sterile pipet
> tip  (attached to the micropipettor or your choice), pick up about half of
> the colony of choice, and resuspend in 25 microliters of ddH20.  I boil for
> 5 minutes, then spin  down 5 minutes to pellet the insoluble
> residue, and use  an aliquot of the supernatant in a PCR reaction, in the
> same way that I would use solubilized DNA.
> 

I was discussing PCR with another grad student, and he thinks you can
amplify genes located on a chromosome by just boiling the cells, pelleting
them, and performing PCR on the supernatant.  I'm very sceptical about
this...surely you'd need to purify the genomic DNA somewhat, especially if
you want high fidelity amplification of your product?  

Curious,
Ed Beaty
Ed_Beaty at qms1.life.uiuc.edu



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