Yeast medium

YPD

Yeast Extract Peptone Dextrose Medium

Makes 1L 10 g yeast extract 20 g peptone 20 g agar (optional, for making plates or slants) 1. Dissolve the above in 900mL ddH2O 2. Autoclave 20 minutes on liquid cycle 3. Add 100 mL of filter-sterilized 20% dextrose. 4. Cool the solution to less than 60 oC before adding zeocin. Note that zeocin in only necessary for selection of Pichia transformants; once transformants are established, zeocin can be omitted from expression studies. 5. Store slants/plates at 4 oC in the dark. Plates are stable for 2 weeks, unless they do not contain zeocin, in which case they are stable indefinitely.

YPDS Agar Plate with Zeocin

Yeast Extract Peptone Dextrose Medium (YPDS) plates containing:

yeast extract 1% (w/v)

peptone 2% (w/v)

dextrose 2% (w/v)
sorbitol 1 M

agar 2% (w/v)

zeocin 400 μg/mL

stored at 4°C

YPDS Agar

Yeast Extract Peptone Dextrose medium with Sorbitol

Makes 1L 10g yeast extract 182.2g sorbitol 20g peptone 20g agar 1. Dissolved the above in 900mL ddH2O 2. Autoclave 20 minutes on liquid cycle 3. Add 100 mL of filter-sterilized 20% dextrose solution 4. Cool to less than 60 0C before adding zeocin. Concentration of zeocin required for selection varies depending on the strain, plasmid, etc. 300 ug/mL is the suggested concentration of zeocin to use for selection, but this may depend on Pichia strain. 5. Store slants/plates at 4oC in the dark. Plates are stable for 2 weeks. (Obiviously, if no antibiotic is included, plates are stable more or less indefinitely.)

YPD (L): yeast extract 10 g, peptone 20 g, glucose 20 g, agar 15 g (if necessary).

BMGY (L): yeast extract 10 g, peptone 20 g, potassium phosphate buffer 0.1 mol, pH 7.0, YNB 13.4 g, biotin 4 · 10–4 g, glycerol 20 g.

Basal salts medium (BSM) (L): H3PO4 26.7 mL, CaSO4 0.93 g, K2SO4 18.2 g, MgSO4·7H2O 14.9 g, KOH 4.13 g, glycerol 40 g, PTM1 4.35 mL,
adjust pH by w = 28–30 % NH4OH to 5.0.

PTM1 (L): CuSO4 · 5H2O 6.0 g, KI 0.08 g, MnSO4 · H2O 3.0 g, Na2MoO4 · 2H2O 0.2 g, H3BO3 0.02 g, ZnSO4 · 7H2O 20 g, FeSO4 · 7H20 65.0 g,
CoCl2 · 6H2O 0.5 g, biotin 0.2 g, H2SO4 5.0 mL.