Help, 5-FOA selection of trxs
ebraun at mail.unm.edu
Tue Jul 11 22:34:59 EST 1995
In article <199507111439.HAA13763 at net.bio.net>
U09668 at UICVM.CC.UIC.EDU (E Li) writes:
> Dear fellow yeasters:
> I have a strain with a genomic disruption of a gene using a URA3 marker, and
> would like to directly transplace the disrupted gene with an intact copy of the
> gene. I was thinking about directly selecting the transformants on FOA
> medium, and was wondering if anyone out there has ever tried this. I don't
> have a selectable marker for the gene I would like to introduce, but I can
> select real transformants as opposed to URA3 revertants, by phenotype.
> Does this approach seem reasonable, and can anyone give me any possible tricks
> or hints about this strategy? Please e-mail me directly with any advice. Thank
> you very much.
> E Li
> Univ. of Ill. at Chicago, Dept. of Biological Sciences
> U09668 at uicvm.uic.edu
I haven't tried doing what you suggest, I thought I bring up a ref.
that you might find useful. In Yeast vol. 11, pp. 53-55 (this is a
couple of months old right now) Fred Winston et. al. describe the
construction of a set of marked strains isogenic to S288C. The first
step was to transform yeast with ura3-52 to 5-FOA resistence. The
procedure used is given in the article. The procedure can also be
found on pp. 309-311 of Meth. in Enzymol. vol. 194 (in Robert S.
Sikorski and Jef D. Boeke "In Vitro Mutagenesis and Plasmid Shuffling:
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