Can't find the gene!

jfh jfhess at ucdavis.edu
Fri Sep 29 15:00:43 EST 1995


Odd.

I have compared bovine and chicken sequences to design primers to
conserved regions and used these primers to clone mouse, human and rat
sequences.

If your reaction works on human genomic DNA, then your problems are with
the mouse DNA in some way.  If you can't get the human DNA to work, then
you need to redo something, either PCR conditions or primers or both.

but,  
First, can you use the primers on cDNA?  The complexity of cDNA is 1/100
that of genomic so you have a better chance to get the real sequence to
amplify.

Second,  if not, don't forget to do southerns on your mouse pcr products.
 always include some human pcr product as a positive for the blot.

If your primers are designed to span an intron, well mouse just may have
a bigger intron.  Some of our work recently has been very frustrating. 
We can amplify cDNA and plasmid sequences but trying to go from genomic
DNA and isolate introns doesn't work. 


John Hess, PhD                    Phone me 916 752 8420
Dept of Human Anatomy             FAX me 916 752 8520
University of Calif               Email me jfhess at ucdavis.edu
Davis, CA                         or leave me alone, your choice.



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