Good morning/evening/afternoon all!
I have one simple question,
i am working with RNA-dependent RNA polymerase, and in its assay was
done as it should be, ive got a ladder of different RNA population,
however i have a feeling that my RNA is stucked up in the wells....
afterwards i realized that my loading buffer does NOT contain
formaldehyde that would react with -N= or :N- or any other possible
target for nucleophilic attack, so the formamide work (denaturing the
RNA at 65C 10 min) is totally useless!
(( And as a result RNA is
stucked up and i dont see a desired product (about 10KB)...
Am I right?
Another prob i have is that the resolution of my gel and its
discreteness is much more to be desired, i use phosphate buffer, 6%
formaldehyde, agarose system [BAD buffer goes hot and it stinks
However, i tried to use MOPS... but it does not contain
formaldehyde... got bad results, really bad..
I have heard about agarose-acrylamide gels, with a fantastic
resolution? Are there?.. and one professor at our fascility says that
it is a real art to make those gels, any comments on that?
Could anyone please suggest me a very NICE approach to solve those
Thank You in advance,