Clonetech DSRed

HENRYT at cellbio.emory.edu HENRYT at cellbio.emory.edu
Wed Nov 24 09:03:30 EST 1999


Hi ,

We've had a similar problem.  At first we thought there was a contaminating plasmid in the original stock sent 
out.  However, it may be some form of plasmid instability:  In our original transformation into E. coli STRAIN 
DH5alpha we got colonies, most of which turned pink/salmon color after a day or so at 37oC grown on LB plates 
with Carbenicillin (50 ug/ml).  I picked a few colonies before they turned pink and streaked them on  grid; these 
were all pink.  After I did a plasmid prep on a culture grown from a single colony  of the original 
transformation and the plasmid was not the DsRed plasmid, as determined by restriction analysis.  I did the same 
digests with the plasmid prep we purchased and saw very light bands when analyzed on agarose gels that appear to 
comigrate with bands I saw in my plasmid isolate.  I took this to mean that there was another plasmid in the 
original stock and I determined it was best to reisolate a single colony from the original transformation.  So, I 
took and restreaked four of the colonies that I had streaked on a grid.  When these grew up I had a mixture of 
colonies:some pink, some white.  Then, I took several single well isolated pink colonies and restreaked them on 
plates and found that I got a mixture of white and pink colonies.  I'm in the process of doing plasmid preps on 
the white and pink colonies to see if they have different size plasmids.  I have informed Clontech of the problem 
and it has headed back to their QC department.  I also would like to know if anybody has had similar problems 
with this plasmid.

Henryt
Henry G. Tomasiewicz, Ph.D.
Emory University
Department of Cell Biology
1648 Pierce Dr.
Atlanta, GA 30322

Ph 404 727-6255
fax 404 727-6256




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