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
cells.
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!
Yiling
------------
Yiling Chen
Department of Botany, Box 355325
University of Washington
Seattle, WA 98195-1800
---
______________O_________oO_____________oO______o_______oO___________________
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
