northern problem - filter is dark (May/01/2003 )
i tried to do northern, my problem is when i exposed the filter(nytran) to the film i got dark picture. i assume that the prob binded unspecificlly but i used salmon sperm, i baked the filter in uv 3 min. each side and i washed the filter 5 times with different concentration of sds and ssc.
any more suggestion why the prob can bind unspecificlly to the filter?
thank is advence
Yuo're not alone!
I think I have the same problem. Even after stripping I cannot get rid of the background. I prehybridized and hybridized according to the protocol and now I still have background.....
Do you mean your whole film was dark? or there was dark stains somewhere in the film?
hi i meant that whole the filter was dark i think i know why. my probe containing a bluescript sequence and i think that it is hybridize some how with the salmon sperm. when i cut the bluescript part of my probe the filter was much less dark.
if u have more suggestions i will glad to hear about them