glass beads for yeast lysis
tomasd at bdg10.niddk.nih.gov
Wed Sep 20 09:06:51 EST 1995
In article <bicfo.14.305E5AC7 at flinders.edu.au>, bicfo at flinders.edu.au wrote:
> hallo all!
> Can someone give me the good oil on using glass beads to lyse yeast? I'm
> 1ml of lysis buffer (50mM HEPES, pH 7.4, 150mM NaCl, 5mM EGTA, 5mM EDTA, 2%
> Triton x-100 and inhibitors) to ~ 6 x 10^8 cells and 1.5g 400 -600u
> acid-washed beads. I'm vortexing them to death - and yet I'm getting zilch in
> the way of lysis! What am I failing to do?
> Replies gratefully received!
> Cheers, Colleen
I use simply 0.05 M Tris/HCl pH 7.5. No detergent, no salt, nothing. I use
about 0.5 g of wet cells in 2 ml breakage buffer. Add beads so that there
is still about 4 mm of free liquid above the beads, and vortex 30''/30''
on ice (10x). At that point they are totally dead, you can barely find a
living cell in micro.
Vidno jak psoj kulky.
(Old Slovak proverb)
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