whole mount in situ--background problem

Hung Fang hfang at fraser.sfu.ca
Mon Apr 19 14:37:13 EST 1993


Hi, there
I am doing whole mount in situ hybridization with sea urchin embryos. I am using DIG labeled probes and using AP-Ab to do the detection. I have some successful experience with coding sequence probe, but I have a lot of trouble when I use in vitro transcribed 3'noncoding sequence as a probe. The problem is although I have tried different things as increasing hybridization stringency, add poly(A) and poly(dA) in the hybridization solution, using different proteinse K digestion condition...I still have the pr




oblem of high background.

Anybody out there has the idea what could be the problem and how to overcome it?Your help is very much appreciated!

hfang at fraser.sfu.ca
or hfang at darwin.biol.sfu.ca

h



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