Yeast cells, PI and cell cycle

From: joan Kalnitsky (jkalnits@vt.edu)
Date: Wed Dec 11 2002 - 12:47:44 EST 

         I have a client who ran some PI cell cycle samples on yeast for
the first time today.  I was expecting a lot of "challenges" , based on
what we read prior to getting started.  Things went well with the exception
of the cell cycle histograms.  I am use to looking at mammalian cells with
very clean distinct peaks, but these peaks, Go/G1, s and G2m were very
broad and not what I am use to.  There were two very distinct peaks with
out much differentiation between them.  When we set the gates based on his
control, the gates appeared to fit the following samples we read, just in a
manner different from what I am use to seeing.
         Does anyone have any feedback on this?  Does yeast give the
characteristic cell cycle histogram?  There were lots and lots of "cells"
in his sample.  Should he try more PI.  His protocol called for RNase
anywhere from 1 hr to overnight.  He used 1 hr, would overnight be better?
         If the histograms looked more familiar to me I would have
confidence in saying we were ready to go, but not having experience in this
area I am a bit hesitant.
         Any and all info will be appreciated.
         Thanks in advance,
         Joan K




         Joan Kalnitsky
         Flow Cytometry Lab Supervisor
         VMRCVM
         (540) 231-4115
         FAX 540-231-7367
         jkalnits@vt.edu

         "It is better to serve than to receive."
            B. Borg

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