Medium data

Search for medium no. = [1148].

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

1148 NITROSOSPHAERA GARGENSIS MEDIUM

KH₂PO₄	`0.05`	g
KCl	`0.075`	g
MgSO₄·7H₂O	`0.05`	g
NaCl	`0.584`	g
CaCO₃	`4.0`	g
Distilled water	`1.0`	L

Mix components and autoclave (CaCO₃ will not fully dissolve). After cooling, add the following solutions to the medium (autoclaved or *filter-sterilized):

50 mM NH₄Cl solution	`20.0`	ml
Trace element solution* (see below)	`1.0`	ml
Selenite-tungstate solution (see Medium No. 431)	`0.5`	ml

Comments: Typically, prepare the medium in glass bottles with plastic caps leaving an ample head space (e.g., 50 ml medium in 250 ml-Duran® glass bottles). Cultivate in the dark without agitation. Check growth by phase-contrast microscopy. Also, it can be checked by measuring ammonium concentration. Once ammonium in the medium is depleted, it can be fed by the addition of another 50 mM NH₄Cl solution (final 1.0 mM). The ammonium-feeding can be repeated several times. If the culture stops the ammonium oxidation or begins to grow even slower, nitrite may be accumulated up to a level causing growth inhibition in the culture (ca. 15 mM). If so, replace the medium as follows: spin down the culture (< 6,500 g for 15 min), remove the supernatant, and add fresh culture medium to the white pellet. Then, suspend the pellet and remove the suspension to a new glass bottle.
Trace element solution:

HCl (conc.)	`2.5`	ml
MnSO₄·5H₂O	`0.048`	g
H₃BO₃	`0.05`	g
ZnCl₂	`0.07`	g
Na₂MoO₄·2H₂O	`0.073`	g
CuCl₂·2H₂O	`0.02`	g
NiCl₂·6H₂O	`0.024`	g
CoCl₂·6H₂O	`0.08`	g
FeSO₄·7H₂O	`1.0`	g
Distilled water	`1.0`	L