Base Layer Agar with Nutrient Overlay Agar

[所属分类:培养基配方] [发布时间:2019-12-6] [发布人:] [阅读次数:] [返回]

山东拓普生物工程有限公司

Shandong Tuopu Biol-Engineering Co.,Ltd

Base Layer Agar with Nutrient Overlay Agar

培养基配方
Composition per 2.5L:
Fat substrate.............................................50.0g
Nutrient agar..............................................1.5L
Basal medium ..............................................1.0L
Fat Substrate:
Composition :
Fat.......................................................50.0g
Preparation of Fat Substrate:  Tributyrin, corn oil, soybean oil,
any cooking oil, lard, tallow, or triglycerides that do not contain anti-
oxidants or other inhibitory substances may be used. Remove free fatty
acids in the fat substrate by dissolving 50.0g of fat substrate in
500.0mL of petroleum ether. Pass the solution through an activated alu-
mina column. Remove the petroleum ether by evaporation on a steam
table under 100% N2 . Autoclave for 30 min at 15 psi pressure–121°C.
Cool to 50°C.
Nutrient Agar:
Composition per liter:
Agar...........................................................15.0g
Pancreatic digest of gelatin....................................5.0g
Beef extract....................................................3.0g
Preparation of Nutrient Agar:  Add components to distilled/de-
ionized water and bring volume to 1.0L. Mix thoroughly. Gently heat
while stirring and bring to boiling. Distribute into tubes or flasks. Au-
toclave for 15 min at 15 psi pressure–121°C. Cool to 45°–50°C.
Source:  The medium is available as a premixed powder from BD Di-
agnostic Systems.
Basal Medium:
Composition per liter:
Agar...........................................................15.0g
Victoria Blue B solution ....................................200.0mL
Preparation of Basal Medium: Add agar to 800.0mL of distilled/
deionized water. If tributyrin is used as the fat substrate, add agar to
1.0L of distilled/deionized water. Autoclave for 15 min at 15 psi pres-
sure–121°C. Cool to 50°C. If tributyrin is not used as the fat substrate,
aseptically add 200.0mL of Victoria Blue B solution. Mix thoroughly.
Victoria Blue B Solution:
Composition per 200.0mL:
Victoria Blue B................................................0.12g
Preparation of Victoria Blue B Solution: Add the Victoria Blue
B to 200.0mL of distilled/deionized water. Mix thoroughly. Filter ster-
ilize. Warm to 50°C.
Preparation of Medium:  Aseptically combine 1.0L of sterile basal
medium with 50.0g of sterile fat substrate in a warm, sterile blender
container. Blend for 1 min until homogenized. Rapidly pour into sterile
Petri dishes in 7.0mL volumes. Dry the surface of the plates by partial-
ly opening the lids in a laminar flow hood for 15 min. Add dilution of
food samples to be tested. When the inoculum is dry, pour nutrient agar
as an overlay onto each plate. Use 10–12mL of nutrient agar per plate.
Use: For the isolation, cultivation, and identification of lipolytic
microorganisms from food.