pCR-Script (Stratagene) cloning kit experience?

Zoltan Penzes PENZES at IRAD.AFRC.AC.UK
Sat Apr 24 17:36:00 EST 1993


Dear Netters,
I tried cloning a 1.4kb PCR product (lots of stuff, clean
band) with  Stratagene's pCR-Script  cloning kit.  I  gel
purified the 1.4kb fragment with GeneClean, then followed
the directions.
I got  27 white  and 270  blue colonies,  far less whites
then the  kit suggests.  More, when  I picked  the  white
ones, only 7 out of 27 remained white after O/N growth. I
tested 24  of the  originally white  colonies and NONE of
them contained  my (any)  insert. The  transformation and
the ligation controls worked, though the results were not
as good as they should have been described in the kit.
Does anyone have any experience with this kit? What could
I do wrong?
 
Thanks for all your comments.
 
************************************
*  Zoltan Penzes
*  penzes at irad.afrc.ac.uk
*  AFRC, Institute for Animal Health
*  Compton, UK
************************************



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