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S. pombe tetrad analysis on minimal media

Jennifer Buslepp buslepp at med.unc.edu
Mon Mar 1 17:37:24 EST 1999

I have a diploid S. pombe strain which is heterozygous for my gene of
interest (also homozygous for  leu 1-32) , where loss of this gene is lethal.  I have
been attempting to isolate mutants of my protein which fail to complement this knockout in
the haploid, using a LEU plasmid containing my gene with random
mutations.  I have isolated several potential mutants using random spore
analysis and am now attempting to verify these by tetrad analysis.   I
have been plating the tetrad spores on pombe minimal media,
and I still do not see any growth after almost two weeks.  I am certain
that the yeast have the plasmid.  My question is whether or not pombe tetrad
 analysis can be done on minimal media, and if not, 
can the analysis be done on rich media without loss of the plasmid?  Any
help with this will be greatly appreciated.

Jennifer Buslepp

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