A rapid technique for the visualization of live immobilized yeast cells.
We present here a simple, rapid, and extremely flexible technique for the immobilization and visualization of growing yeast cells by epifluorescence microscopy. The technique is equally suited for visualization of static yeast populations, or time courses experiments up to ten hours in length. My microscopy investigates epigenetic inheritance at the silent mating loci in S. cerevisiae. There are two silent mating loci, HML and HMR, which are normally not expressed as they are packaged in heterochromatin. In the sir1 mutant background silencing is weakened such that each locus can either be in the expressed or silenced epigenetic state, so in the population as a whole there is a mix of cells of different epigenetic states for both HML and HMR. My microscopy demonstrated that there is no relationship between the epigenetic state of HML and HMR in an individual cell. sir1 cells stochastically switch epigenetic states, establishing silencing at a previously expressed locus or expressing a previously silenced locus. My time course microscopy tracked individual sir1 cells and their offspring to score the frequency of each of the four possible epigenetic switches, and thus the stability of each of the epigenetic states in sir1 cells. See also Xu et al., Mol. Cell 2006.
Zawadzki
Karl
K
Department of Molecular Biology, Princeton University, USA. kzawadzk@princeton.edu
Broach
James
J
D018914:Cells, Immobilized / Q000166:cytology*; D002874:Chromosome Mapping; D044127:Epigenesis, Genetic; D020868:Gene Silencing; D008856:Microscopy, Fluorescence; D018715:Microscopy, Video; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000235:genetics