cell sorting live yeast
lweisman at BLUE.WEEG.UIOWA.EDU
Mon Jan 13 10:24:49 EST 1997
We frequently sort live yeast and developed a FACS screen for mutants
defective in vacuole inheritance. Details of our protocol are described
in Wang et al. MBC 7:1375-1389 (1996). The main complication for sorting
yeast is that individuals are much more variable in light scattering
measurements, both foward angle light scatter and orthogonal light
scatter. (This is probably due to 1) actual differences in cell size and
shape, especially in a mutant population 2) unbudded vs large budded cells
and 3) some clumps instead of single cells passing through the detector.)
So when we sort, we general display both fluorescence and foward angle
light scatter on a log scale. If you have further questions, you may
contact me by email. Hope this helps.
On Thu, 9 Jan 1997, Bev Rothermel wrote:
> I'm looking for information on flow cytometry cell sorting of live
> unfixed yeast cells. Will it work, and does anyone have a good method?
> I want to use GFP as the cell marker and be able to grow up the cells
> after sorting for multiple rounds. The only methods I've seen call for
> fixing the cells prior to FACS.
> Bev Rothermel
> rothermel at ryburn.swmed.edu
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