help in doing yeast transformations !!!

ed s535290 at aix1.uottawa.ca
Sun May 4 16:51:29 EST 1997


Hi all,

Our electroporator is on the fritz, and I need to do yeast transformations
preferably using the LiAc method. I have a somewhat modified (and
much simpler !!!) version from the one that appears on the red book.
Nonetheless, I read the red book for pointers...the thing that struck me
was that it mentions that single most critical parameter in transformation
efficiency when using the LiAc method is the quality of the carrier DNA
being used...in particular, that it be totally denatured and of a median
fragment size of about 7 Kb (!?!?!?). The question I ask of anyone who's
done yeast transformations, is whether you prepare your own carrier DNA or
purchase it from somewhere ?  It mentions sonication in the shearing of
the DNA, but without access to a sonicator, I don't know how to go about
getting the fragment sizes right...in the red book they even suggest
running an aliquot to check median fragment size...are they being
unnecessarily anal retentive about this or should I definitely be
concerned about this ? I don't need an incredible transformation
efficiency, as long as I have a few different transformants...please share
your knowledge with me :)

thanks in advance,

Ed




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