Cracking Assay for Recombinant Clones

Matthew L. Brown, Ph.D. gemini at
Fri Jun 20 12:11:05 EST 1997

Fellow Molecular Biologists,
	Some of you may already do this so I'm sure that this is not a "novel"
idea but I though others might appreciate the suggestion. I routinely
perform "cracking" assays to identify recombinant clones after
transformation. I have always used a slight modification (lower volumes)
of the method detailed in Sambrook et al. (Blue Books). I make up stocks
of the reagents except the lysis buffer which I make up fresh each time.
This procedure works pretty well. I also use Qiagen spin preps to do my
minipreps once I have identified interesting clones (this is not an
advertisement so _please_ no flaming!). Anyway, whenever I finish a
Qiagen kit there is always a substantial amount of the Suspension
buffer, lysis buffer, and neutralization buffer. In the past I had
simply thrown these away. Recently I began saving these buffers and
"recycling" them by using them in my Cracking Assays. It works great!
Saves me time and since the resuspension buffer contains RNase it
removes the annoying RNA band that had always been present in my
cracking gels. I don't know why it took so long for this thought to
occur to me. Anyway, like I said this is not novel and not Earth
shattering but thought some people might benefit from the suggestion.
Good Luck,

Matthew L. Brown, Ph.D.
Research Fellow
Department of Medicine
University of Alabama at Birmingham
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