Dear yeast enthousiasts,
I have a question concerning "synthetic viability".
We want to identify novel proteins that interact with a known protein. We
have two mutants in this protein that cause a non-conditional lethal
phenotype, i.e. cells cannot grow on these mutant proteins under any
We want to isolate suppressor mutations and we came up with the following
concept: we want to construct a yeast strain that is disrupted in its
chromosomal locus and carries two plasmids, one with the wild-type gene on
a URA3 plasmid and one carrying the mutated gene on a LEU2 plasmid.
Normally, these cells cannot lose the URA3 plasmid and should therefore be
unable to grow on 5-FOA. After mutagenesis, we want to screen for cells
that can grow on 5-FOA with the expectation that these cells contain a
(suppressor) mutation that allows the cells to use the other, mutated
protein. It is a bit reminiscent of a "reverse synthetic lethal" screen.
Obvious artefacts that we can think of is loss of a functional URA3 plasmid
marker, but this should easily be screened out.
My questions to you, the yeast community, are:
(i) has this been done before (presumably yes...) and does anybody have
information about this?
(ii) is there a simple(r) alternative technique that we have overlooked?
(iii) does anybody have comments on the feasibility of this approach, both
theoretically and in practice (background artefacts)?
Many thanks for your help, I would be willing to put a summary on the net
if people are interested.
email venema at chem.vu.nl