PCR from yeast genomic DNA
vvsvetlov at utmem1.utmem.edu
Sat Jan 13 01:18:41 EST 1996
In article <lichten-0901961049190001 at 22.214.171.124>,
lichten at helix.nih.gov (Michael Lichten) wrote:
> In article <4cpqaj$2a2 at no-names.nerdc.ufl.edu>, Jon Stewart
> <jds2 at ufcc.ufl.edu> wrote:
> > I need to PCR amplify two Saccharomyces genes of approx. 1000 bp,
> > neither of which contains an intron according to the sequence database.
> > Since I have never attempted PCR from yeast genomic DNA, I have some
> > questions.
You can PCR yeast directly from colony or isolate DNA from liquid culture
by rapid release methods - protocols available in CPMB and in White yeast
methods book. I take 5-20 ug of genomic DNA per 100 ul reaction with Pwo.
Rapid cycling (5-10 sec each step) works the best - cleaner bands, no
non-specific amplifications. There are a lot of people working with yeast
around you, it should be easy to get some DNA or cells. If you can't get
it, send me your Fedex number - I'll send you some DNA from relatively WT
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