Synchronising yeast cells

Hernan Espinoza espinoza at cgl.ucsf.edu
Wed Oct 23 11:23:35 EST 1996


karthi at deakin.edu.au (Karthi Ramachandran) writes:

>1. My strains are Mat alpha. Is a-factor commercially available or do I have 
>to allow them to switch mating types to Mat a and use alpha factor instead.

	a-factor is not commercially available to my knowledge.  Theoretically,
you could purify a-factor from a-cell conditioned media, but switching mating
types would be much easier.

>2. Do you have a good protocol for synchronising yeast cells. I need to know 
>the concentration of the mating harmones used.

	Treat MATa cells with 1 microgram per ml alpha-factor (1000X stock =
1 mg/ml in DMSO) for 90 min.   Check cells under the microscope for uniform
shmooing.  Wash twice[QUICKLY!] with prewarmed 30C media.  They're off.
We take samples every 10-20 min for 2 hrs depending on the particular
experiment.  You can test the synchrony of the cells by looking at the
budding index (# budded cells/total cells) over the time course.

>3. Our local peptide synthesise facility wants A$ 1500 for 20mg of the alpha 
>factor peptide. Is that the normal price or am I being ripped off?

	The same amount from Sigma would cost ~US$ 1300.  You'll have to do 
the math since I don't know the conversion factor.  Actually,  there are
probably cheaper sources out there, but I don't know them.  Anyone?

		-Hernan



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