Developmental Northern

martin martinNOmaSPAM at biologie.uni-freiburg.de.invalid
Tue Jan 25 12:42:55 EST 2000


Hi everybody,

even though my DIG-Northerns work pretty well, I have a problem with my
loading standard. I use a DNA-DIG-rp49-probe, which (the literature
says)
is a constitutively expressed gene and has been used as a loading
control
for developmental northerns in other papers. I check my RNAs (total) in
a
photometer and am pretty accurate in respect to quantity while loading
the
sample on the gel.  The problem is, that rp49 always gives me totally
different signal strengths after hybridization for the different
developmental stages. Embryo and L1 is strong and the rest pretty weak.
Unfortunately, especially those are weak that also do not show
expression
The RNAs do not seem to be degraded, since I see partially weak, but
always
distinct rp49 signals for all stages and no smear.

What should I do? Is the photometer the reason? I do not look at the
gel,
since EtBr seems to make a pretty big background. Is Rp49 the reason.
What other method could I use to quantitate the RNA?

Thank you in advance!!

Martin


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