Help Needed for Oocyte RNA Expression Study

[wch190 at lulu.acns.nwu.edu]

In article <3ar1or$6r5 at newsbf01.news.aol.com>, riversloth at aol.com
(RiverSloth) wrote:

> I am about to send some in vitro transcribed RNA to collaborators at
> another location where they will microinject the RNA into Xenopus oocytes.
> We hope to get the RNA translation product incorporated into the oocyte
> cell for receptor binding assays-the RNA codes for a neuropeptide hormone
> receptor.
> 
> Since oocyte expression studies are a new area of study for me, I was
> hoping to get some advice on a couple of questions I have:
> 
> 1.  Will the length of the 5' untranslated region of the RNA affect the
> translatibility of injected RNA in the oocyte system?
> 
> 2.  Will spurious AUG codons occuring 5' of the "correct" AUG codon for
> the receptor protein open reading frame prevent translation of the correct
> open reading frame, particularly if the spurious AUGs have termination
> codons? Or will the ribosomes resume scanning the RNA for the next AUG,
> eventually reaching the receptor open reading frame?
> 
> I appreciate any help you can give me. I am trying to anticipate problems
> before they arise. I expect any problems with the assay will be laid at
> the feet of the RNA supplier (me!) so I hope to be ready with some
> answers.
> 
> Thanks,
> 
> Felix D. Guerrero, PhD
> USDA-ARS
> Kerrville, TX

The length of the 5' untranslated will affect the stability of the RNA ..
it would be better if you have a poly A tail at the end 

As for the AUG I think the first AUG is taken as the start 

Venita De Almeida 
Northwestern Univ 
Evanston Ill