Dear Yeast People,
I use a PCR URA3 linear cassette to disrupt an essential gene in diploid
yeast. This yeast strain has a point mutation in URA3 gene and turns out
a URA- background. The disruption of this gene is lethal in haploid yeast
Theoretically, I should at least get some transformants which remain only
one wild type allele. I have been working on both Lithium Acetate and
electroporation transformation methods for a while, but these methods never
bring me the right transformants. I always get few transformants with the
homologous wild type allele but never get the heterozygous one with one
wild type and one marker exchanged allele.
So a question comes out of my mind. Is it possible that the phenotype is
lethal even with a wild type allele in diploid yeast cells? Could any one
suggest me some relevant references in this case?
So many thanks!
Department of Botany, Box 355325
University of Washington
Seattle, WA 98195-1800
YEAST bionet newsgroup see: http://www.bio.net/hypermail/YEAST/
YEAST e-mail: messages sent to yeast at net.bio.net
subscribe: e-mail biosci-server at net.bio.net with: subscribe yeast
unsubscribe: e-mail biosci-server at net.bio.net with: unsubscribe yeast
YEAST on the WWW: http://genome-www.stanford.edu/Saccharomyces/VL-yeast.html
problems with the YEAST newsgroup? E-mail the moderator: francis at cmmt.ubc.ca