RNA Slot-blotting woes

Gabriel Romero ez008413 at dale.ucdavis.edu
Tue Jun 4 17:45:14 EST 1996


I'm trying to quantify my gene's transcript in several tissues using 
slot-blotting of total RNA and known amounts of target DNA as standards. 
My previous northerns show comparable high level expression let's say in 
tissues a,b and c. When I slot-blotted the same amount of total RNA from the 3 
tissues as I ran for the northrens, I got starkly different results: 
such as, only tissue a showed signal with b and c showing nothing. In short, 
the northern data and slot-blot data are inconsistent.  Because my northerns 
have been reproducible, I suspect something is wrong with the 
slot-blotting. So, in my subsequent attempts, I've turned down the vacuum 
and also added carrier tRNA to improve RNA retention on 
the blot. Still no improvement. Could somebody help me troubleshoot?  I'd 
appreciate any advice.




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