Plate pouring

To make plates, it is the easiest to prepare 2 L batches. Make one 2L flask with 1L of water and 40g of agar. Take a 1L bottle and use that for the actual medium (so use 2 times the amount for 1L). Make it up in 900 ml of water, to leave room for 100 ml of 40% glucose. Put a stirbar in the bottle. Autoclave the flask and the bottle, and pour the contents of the bottle into the flask. Then add glucose, amino acids etc.

YEPD

For 1L medium:

• 20g yeast peptone
• 10g yeast extract
  ----autoclave----
• add 50 ml 40% glucose

Synthetic Complete (SC) -Leu -Trp -His -Ura

For 1L medium:

• 1.3g AA powder
• 1.7g YNB (without AA and Ammonium Sulfate)
• 5g Ammonium Sulfate
• Adjust pH to 5.9
  ----autoclave----


• add 50 ml 40% glucose
• add 8 ml of each additional AA you want (Leu, Trp, His, Ura) and 3AT or 5-FOA

SC additives

100 mM Histidine (don't autoclave)
100 mM Leucine
40 mM Tryptophan (keep in aluminum foil at 4°C, don't autoclave)
20 mM Uracil

20 mM 3AT = 1.68 g/L
5-FOA = 1g/L or 2g/L
Cycloheximide = 1µg/ml final from a stock of 10mg/ml in ethanol

The color codes for on the plates

Amino Acid powder

Mix 3g each of the following Sigma amino acids. The mixture will remain stable for up to 6 months.