PCR directly from bacterial colonies

Hecht Hecht at mars.rowan.edu
Mon Aug 14 19:41:39 EST 1995


I have also carried out PCR routinely from bacterial colonies, although
my experience is in cloning products amplified from the chromosome.  I
set up my PCR reaction sans oil and polymerase, pick a colony using a
clean sterile pipetman tip, and scrape the colony off into the reaction
tube.  Boil the reaction tube for 5 minutes.  Spin down to collect the
drops, add polymerase, cover with oil, and you're ready to start the
thermal cycler.  
     A couple of notes:
     (1)  Since we re-use toothpicks in our lab, we prefer to pick the
colonies using the pipetman tips [yes, we autoclave our toothpicks
before re-using them ;=) but you just never know, eh?].  
     (2)  There are a number of protocols floating around out there for
this technique, some of which claim that colony PCR is more less
forgiving than when you use purified DNA as your source; in my
experience, the same reaction conditions will work when using purified
DNA or a boiled colony.



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