Medium data

Search for medium no. = [862].

Unless otherwise stated, sterilize media by autoclaving at 121°C for 15 min.

862 STEROIDOBACTER MEDIUM

NaCl	`1.0`	g
MgCl₂·6H₂O	`0.4`	g
KH₂PO₄	`0.2`	g
KCl	`0.5`	g
NH₄Cl	`0.25`	g
CaCl₂·2H₂O	`0.15`	g
Na₂SO₄	`0.07`	g
NaNO₃	`0.42`	g
Distilled water	`1.0`	L

For growth with testosterone, dissolve testosterone in acetone (20 mg/ml) and dispense portions of the solution, e.g., 0.1 ml portions for 10 ml medium, to anaerobic culture tubes. Let the solvent evaporate to dryness. Dispense 10 ml portions of medium in the culture tubes and stream with N₂-CO₂ (80:20, v/v). Close the vessels and autoclave. Treat the vessels in an ultrasonic bath to detach and suspend the testosterone.
For growth with heptanoate, omit testosterone. Sterilize the gassed medium in anaerobic vessels by autoclaving. Add 2.5 mM heptanoate from a 20-fold filter-sterilized anaerobic stock solution after cooling.
After autoclaving, add (per 10 ml) 0.3 ml NaHCO₃ solution (84 g/L, autoclaved under a CO₂ atmosphere), 1 µl Trace element solution SL-10 (see Medium No. 433), 1 µl Selenite-tungstate solution (see below) and 1 µl Vitamin solution (see below). Adjust pH to 7.2. Incubate the cultures in the dark for up to 4 weeks. Briefly shake the cultures once per day.
Selenite-tungstate solution:

NaOH	`0.5`	g
Na₂SeO₃·5H₂O	`3.0`	mg
Na₂WO₄·2H₂O	`4.0`	mg
Distilled water	`1.0`	L

Vitamin solution:

Vitamin B₁₂	`50.0`	mg
Pantothenic acid	`50.0`	mg
Riboflavin	`50.0`	mg
Pyridoxamine·HCl	`10.0`	mg
Biotin	`20.0`	mg
Folic acid	`20.0`	mg
Nicotinic acid	`25.0`	mg
Nicotinamide	`25.0`	mg
α-Lipoic acid	`50.0`	mg
p-Aminobenzoic acid	`50.0`	mg
Thiamine·HCl·2H₂O	`50.0`	mg
Distilled water	`1.0`	L