FOA selection in pombe

Wenyi Feng wfeng at newssun.med.miami.edu
Thu Feb 8 15:43:13 EST 2001


Dear yeast community,
    I have been using 5'FOA selection for ura- strain in pombe by
transforming cells with a linear fragment of an epitope tagged gene X
flanked by its genomic 5' and 3' UTR sequences.  I'm hoping to replace
the ura4+ gene that has been placed in  the gene X locus previously with
this epitope tagged gene X through double crossover of homologous
recombination, thus yielding the ura- progeny.  So far I have no success
whatsoever, i.e. no transformants at all when selected on 0.1% 5'FOA
plates.  I was told that cell viability on FOA plates is low, question
is how do I circumvent this problem.  Does anyone know what is the
frequency of obtaining a positive clone through such a transformation?
Any suggestions?  Please help.

Thanks,
Wenyi Feng


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