Plates for sporulation.
Kac 20 gm
yeast extract 2.2 gm
dextrose 0.5 gm
complete mixture 870 mg <-contains all the amino acids
agar 25 gm
H20 1000ml
pH to7.0
I have not heard of a problem with ura- strains, we use them all the time.
s288c is not the best sporulator in the world, but you should see some. how
long are you leaving your cells on sporulation media?
if the above recipie does not work then try
20g Kac
25g agar
1000ml H2O
pH 7.0
we call it minimul sporulation media ( I guess you could supliment with a
spot of ura) and it shocks most things into spores!
I assume you are selecting for diploids in each case by streaking for
singles on synthetic compleat media lacking any uracil, then patching out a
colony onto yepd to get good strong growing cells.
I am developing a web site with all the tricks of the trade from our yeast
lab at oxford, but it may be a while before a public release. if anyone has
any ideas they would like to see then E-mail me.
My real e-mail is
aweather at molbiol.ox.ac.uk
but I will get stuff if you just reply.
Hope that helps
Aidan
"Yeast Laboratory 1" <Yeast1 at demogr.mpg.de> wrote in message
news:80rpjs$6fq at net.bio.net...
> Hi everyone,
>> I am having a problem with sporulating my cells. They simply do not make
> spores. They two strains that I have tried to sporulate look like
diploids
> under the microscope (they are larger than haploids and they bud from each
> pole). The genotypes of the strains are:
>> Strain one: Mat a/Mat alpha; ura3-52/ura3-52; trp1delta63/TRP1;
> leu2delta1/LEU2; his3delta300;HIS3; GAL2/GAL2 (S288C background)
>> Strain two: Mat a/Mat alpha; ura3-52/ura3-52 (S288C background)
>> Unless there is some problems with sporulation in a ura3-52, I suspect it
> could be the type of sporulation media or even the enzyme (10%
> Beta-glucuronidase). Has anyone else encounted this problem?
>> Thank you in advance for your help!
> Christi
>gendron at demogr.mpg.de
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