probe labelling for non-radioactive in situ hybridization

Stephen Saul saul at hawaii.edu
Mon Mar 17 19:00:20 EST 1997


Dear netters:
	According to the in situ hybridization protocol I have for flies, the
labelled probe concentration should be 20 ng/ul or higher. 
Unforturenately, after plasmid digestion, gel extraction, and Boehringer's
DIG DNA labelling kit labelling, the probe concentration was only 0.5 ng/ul
(1/10 of the control probe in the kit).  I did not get any brown color on
the chromosome by using this probe.  
	I have contacted with the manufacturer and was suggested to use proteinase
K to digest the restrictive enzyme which may affect the labelling reaction.
 It did not help.  
	Please help me with your experience.
	Thank you.

					Jingwei 



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