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lysis of saccharomyces

Alex Brands abbrands at artsci.wustl.edu
Fri Sep 1 12:07:40 EST 2000


On 30 Aug 2000, Anjali Prashar wrote:

> Would you know of a quick, cheap and easy way of lysing Saccharomyces 
> cerevisiae cells
> 
> Anjali

Here is the protocol I use to break open yeast cells for the purpose of
recovering plasmids:

-Spin down 1.5mls overnight culture
-resuspend in 100ul:
	2.5M LiCl
	50mM Tris-HCl pH 8.0
	4% Triton X-100
	62.5mM EDTA
-Add 150ul phenol/chloroform
-Add 200mg glass beads (~500 micron)
-Vortex 2 minutes
-Spin 5 minutes
-Transfer upper aqueous phase to a new tube
-Extract with phenol/chloroform
-Spin 5 minutes
-Transfer upper aqueous phase to a new tube
-precipitate with 2.5 volumes EtOH, on ice for 10 minutes
-spin 15 minutes at 4C
-Wash pellet with 300ul 70% EtOH and spin 2 minutes
-Dry pellet and resuspend


If you are doing this to recover proteins or RNA, you will need to do
something to stop protease or RNase activity.

Alex


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