Preparation of Isotopically-Labeled Filamentous Virus

The protocol is the same as for non-labeled virus, except a defined (minimal) medium is used for the growth of the virus. The recipe for this medium depends on the label that is being incorporated.

1. Defined Medium

Min Salts 11.3 g Na2HPO4 3.0 g KH2PO4 0.5 g NaCl 4.0 g NH4Cl 100 mL H2O adjust pH to 7.5 using NaOH	Thiamine 40 mg in 10 mL H2O
	CaCl2 40 mg in 10 mL H2O
Glucose 10 g in 35 mL H2O	MgSO4 0.25 g in 10 mL H2O
Labeled Amino Acid 11-15 mg of labeled amino acid per residue present in protein in 20 mL H2O	L-Amino Acids 50 mg Val 50 mg Leu no unlabeled L-amino acid of the amino acid that is being labeled 11 mg of all other L-amino acids 825 mL H2O

Autoclave all solutions separately.


2. For ON

34 mL L-aa
5 mL min salts
2.5 mL glucose
1.0 mL labeled aa
0.5 mL MgSO₄
0.5 mL CaCl₂
0.5 mL thiamine

Aseptically, add the appropriate amount of each solution to a small, sterile flask.


3. For Large Scale Preparation (856 mL)

Aseptically, pour all solutions into L-aa solution.

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