In article <JOHNS.SIMON-1507961629160001 at 127-90.med.utah.edu>,
JOHNS.SIMON at HSC.UTAH.EDU (laurie JACKSON) wrote:
> i have a strain that i need to transform with a library at high
> transformation efficiency but this mutant strain does not transform well
> at all. This strain must also carry a selection plasmid at the same time
> and be grown on multiple selection media. Any ideas?
There's lots of reasons that yeast might not transform well. I've
found that with a lot of mutants, there is a cell wall defect that makes
them very osmo-sensitive when placed in Lithium acetate. If this might be
the case, try adding 1.2M sorbitol in with your competance buffer. You
may also want to try electroporation -- efficiency is quite dependent on
the strain background, but it can increase efficiency 100 fold. Other
things to vary would be the length of the heat shock. For a good
reference on efficiency look at Geitz et. al., (1995) YEAST, 11: 355-360.