Excellent method to stain DNA in yeasts for cell cycle analysis is described by Steven Reed in CELL CYCLE. Below I am providing the link to CELL CYCLE (which still is available online). His atricle in in issue # 2. Dr. Reed also prepared a protocol on this method which is being now included in Current Protocols in Cytometry. Zbigniew Darzynkiewicz, M.D., Ph.D. Director, Brander Cancer Research Institute New York Medical College 19 Bradhurst Ave. Hawthorne, NY 10532 tel: 914-347-2801 fax: 914-347-2804 http://www.geocities.com/z_darzynkiewicz http://www.landesbioscience.com/journals/cellcycle.html > -----Original Message----- > From: joan Kalnitsky [SMTP:jkalnits@vt.edu] > Sent: Wednesday, December 11, 2002 12:48 PM > To: Cytometry Mailing List > Subject: Yeast cells, PI and cell cycle > > > 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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