MEDIUM FOR EMBRYO RESCUE (FOR 1 L)

1. Put 500 mL of distilled water in a 1-L beaker.

2. Add the following:

• ucrose (6 % final concentration) – 30 g

• i-inositol (100 mg/L stock is 0.5 g/50 mL) – 10 mL

• Thiamine (0.4 g/l stock is 2 g/50 mL) – 10 mL

• Amino acids L-tyrosine, L-argenine, glycine – 10 mL (10 mg/L stock is 50 mg each aa/50 mL H₂O)

• Kinetin (for very small enbryos only) – 2 mg

• Muraschige–Skoog Organic medium – 34.6 g

• NaH2PO4H2O (stock 17 g/L) – 10-mL stock

• Inorganic salts

• Organic constituents

Dilute to 800 mL.

Adjust pH to 5.7 with NaOH..

Bring volume to 1,000 mL.

Add 8 grams Bacto® agar.

Melt agar in microwave for 3 minutes or dissolve while stirring on a hotplate.

Pour ~20 mL into Kimax culture tubes (50 mL) and cap.

Sterilize by autoclaving for 15 minutes.

Slant in tubes or pour into petri dishes.