nonradioactive screening

Dr. Guy Hoelzer hoelzer at unr.edu
Wed Jul 13 11:47:46 EST 1994


In article <2vuuqa$6bv at nntp2.Stanford.EDU>, btoomey at leland.Stanford.EDU
(Barbara Holland Toomey) wrote:
> 
> I getting ready to screen a cDNA library with a DNA probe. I am interested
> in nonradioactive methods to label this probe (I have a 200bp PCR product) 
> since I would like to have kids someday. I am considering Amersham's ECL 
> 3'-oligolabelling system, but I have no experience with any of the 
> nonradioactive methods. I would greatly appreciate any advice on this kit or 
> any others that might be better. Thanks!!!
> Barbara Toomey

I agree with Emma that a 200 bp fragment may be too small for random-primer
labelling.  I have had success with the Genius Kit from Boehringer
Mannheim.  With a 200 bp fragment, you can use the tailing reaction to
create a very sensitive probe.  Using this kit, I have probed a library
with an oligo that was labelled on the 3' end with a single dig-dUTP.  This
yielded sufficient sensitivity.

******************************************************************************
Guy Hoelzer                                                  
hoelzer at unr.edu
Dept. of Biology
University of Nevada Reno
Reno, NV  89557
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