Smolke:Protocols/Yeast Media Recipes: Difference between revisions
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*Add 200mg/L for maintenance (in liquid culture media) | *Add 200mg/L for maintenance (in liquid culture media) | ||
'''For yeast | '''For yeast counterselection:''' | ||
*Add 1 g/l of 5-FOA to YNB-SC solid media to | *Add 1 g/l of 5-FOA to YNB-SC solid media to counterselect for URA3 | ||
**While adding 20% dextrose and 10x SC AA mix, have 5-FOA dissolved in DMSO and add before stirring and pouring out agar (for a typical single sleeve 500ml media batch, this can be 500mg of 5-FOA in 5ml DMSO) | **While adding 20% dextrose and 10x SC AA mix, have 5-FOA dissolved in DMSO and add before stirring and pouring out agar (for a typical single sleeve 500ml media batch, this can be 500mg of 5-FOA in 5ml DMSO) | ||
*Add 1.3 g/l of 5-fluorocytosine (5-FC) to YNB-SC solid media to counterselect for FCY1 | |||
**dissolve 2.6 grams into 200 ml, filter sterilize, add 50ml per 500ml sleeve of plates | |||
**if stored cold or frozen, will crash out of solution; heating helps dissolve | |||
**detailed useful compound info here: http://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma/Product_Information_Sheet/f7129pis.pdf | |||
*For alpha-AAD/LYS2,5 counterselection replace nitrogen source with 2g/L (0.2%) D,L-alpha-aminoadipate | |||
**dissolve 1 gram alpha-AAD in 20ml of water and pH with KOH to pH 6.0 (otherwise insoluble), filter sterilize and add to YNB with special AA mix | |||
**two recipes have been shown to work for this counterselection | |||
***YNB-alpha-AAD + URA,TRP,HIS,LEU,LYS | |||
***YNB-alpha-AAD + URA,TRP,HIS,LEU,LYS,MET,ADE | |||
***special media informed by Gottschling lab site: http://labs.fhcrc.org/gottschling/Yeast%20Protocols/lore.html | |||
-'''[[User:Michael S. Siddiqui|Michael S. Siddiqui]] 21:53, 4 January 2011 (EST)'''* 01/04/2011 ( | -'''[[User:Michael S. Siddiqui|Michael S. Siddiqui]] 21:53, 4 January 2011 (EST)'''* 01/04/2011 (aided by [[User:Andrewb|Andrew Babiskin]]) |
Revision as of 14:43, 18 March 2014
Yeast Nitrogen Base Minus Amino Acids (YNB-AA) Recipe
168355)
Autoclave (store at RT for media stocks) To use as working media:
Yeast Nitrogen Base Minus Amino Acids Non-Inducing, Non-Repressing (YNB-AA NINR) Recipe
To use as working media:
Yeast Peptone (YP) Media Recipe
Autoclave (store at RT for media stocks) To use as working media: After autoclaving add:
To pour any of these media stocks as plates:
For yeast KanMX (G418) selection:
For yeast counterselection:
-Michael S. Siddiqui 21:53, 4 January 2011 (EST)* 01/04/2011 (aided by Andrew Babiskin) |