This is a cached page for the URL (http://hdklab.wustl.edu/lab_manual/yeast/yeast11.html). To see the most recent version of this page, please click here.
Protocol Online is not affiliated with the authors of this page nor responsible for its content.
About Cache

Yeast Media, Solutions and Stocks

Updated Jan. 17, 1991

Jim Howe and C. Helms


Yeast Media:

AHC Medium (a selective medium for growing yeast strain AB1380, a host for YACs)

AHC plates

SD Medium (a synthetic, minimal medium)

SD plates

Sorbitol-Agar Medium (1 liter)

Sorbitol-Agar Medium Top Agar

Sporulation Medium Plates

1% potassium acetate10 g
0.1% yeast extract1 g
0.05% glucose0.5 g
2% agar20 g
distilled water1000 ml

YPD Medium (an enriched, non-selective yeast growth medium)

  1. 10 g yeast extract
  2. 20 g Peptone
  3. 1 liter dH2O
  4. adjust pH to 5.8 with approx. 50 l 12 N HCl
  5. autoclave 30-45 minutes and cool to 65 degrees C, then add 50 ml of sterile 40% glucose

YPD plates

Synthetic Media Supplements

Stock solutionml stock for
ConstituentFinal mg/Lper 100 ml dH2O1 liter media
adenine sulfate20200 mg*10
uracil20200 mg*10
L-tryptophan201 g2
L-histidine-HCL201 g2
L-arginine-HCL401 g4
L-methionine201 g2
L-tyrosine50200 mg25
L-leucine601 g6
L-isoleucine601 g6
L-lycine-HCL501 g5
L-phenylalanine501 g*5
L-aspartic1001 mg10
L-glutamic acid1001 g*10
L-valine1503 g5
L-threonine2004 g*5
L-serine4008 g5
* Store at room temperature

Solutions and stocks:

40 mM EDTA, 90 mM 2-Mercaptoethanol (2-ME)

40 ml0.5M EDTA, pH 8.0
3.8 ml2-Mercaptoethanol (12M stock)
456.2 mlsterile ddH2O
500 mlStore at room temperature.

5X HPB (High Phosphate Buffer) (for 1 liter)

  1. 146.1 g NaCl
  2. 134.0 g Na2HPO4
  3. 9.3 g Na2EDTA
  4. pH to 7.0 with concentrated HCl
  5. Autoclave to sterilize
SCE (100 ml)

Final concentration
2M sorbitol50 ml1.0 M
1M sodium citrate10 ml0.1 M
0.25M EDTA, pH7.024 ml60 mM
sterile ddH2O16 ml
------
100 ml

SCE/ DTT/ Lyticase (for 40 ml, prepare fresh each time):

  1. 40 ml SCE
  2. 300 l 2 M DTT
  3. 5600 units Lyticase

SCE/2-ME/Lyticase (2 ml)

  1. 2 ml SCE
  2. 16 l 2-ME
  3. 0.2 mg Lyticase

Mbo I Buffer

  1. 100 mM NaCl
  2. 10 mM Tris-HCl, pH 7.4
  3. 10 mM MgCl2
  4. 1 mM Dithiothreitol
  5. 100 g/ml bovine serum albumin

Miniprep Lysis Buffer

Final concentration
1M Tris-HCl, pH 7.45 ml50 mM
0.5M EDTA, pH8.05 ml25 mM
5M NaCl10 ml500 mM
1M MgCl2300 l3 mM
12M 2-Mercaptoethanol25 l3 mM
Nonidet P-40100 l0.1 %
10% SDS10 ml1.0 %

Large Scale Prep Lysis Buffer

0.5 M Tris-HCl, pH 9.02.5 ml 2 M Tris-HCl stock
3% sarkosyl 3.3 ml10% sarkosyl stock
0.2 M EDTA4 ml 0.5 M EDTA stock

1 mM Tris/1% Sarkosyl (for 1 liter)

  1. 1 ml 1M Tris-HCl, pH 7.5
  2. 100 ml 10% sarkosyl
  3. 899 ml dH2O

200 mM Tris/2X SSC (1liter)

  1. 200 ml 1M Tris-HCl, pH 7.5
  2. 100 ml 20 X SSC
  3. 700 ml dH20

Yeast prehyb/hyb solution (for 10 ml)

  1. 7 ml sterile dH2O
  2. 2 ml 5X HPB
  3. 1 ml 10% sarkosyl

Yeast Lysis Buffer

  1. 50 mM EDTA
  2. 1% sarkosyl
  3. 10 mM Tris-HCl, pH 8.0
  4. 1 mg/ml proteinase K

References:

Sherman, F., Fink, G. R., and J. B. Hicks. (1986). Methods in Yeast Genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.

Brownstein, B. H., Silverman, G.A., Little, R.D., Burke, D. T., Korsmeyer, S. J., Schlessinger, D., and M. V. Olson, (1989) "Isolation of single-copy human genes from a library of Yeast Artificial Chromosome clones". Science 244: 1348-1351.