Fast plate method for screening of methionine producing bacteria

A plate and plate culture medium technology, applied in biochemical equipment and methods, microbiological determination/inspection, etc., can solve problems such as time-consuming, interference with methionine content determination, etc.

Inactive Publication Date: 2019-09-17
南京先华生物科技有限公司
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Problems solved by technology

[0005] The determination of methionine content in fermentation broth is mostly determined by amino acid analyzer. This method is more accurate, but the disadvantage is that it takes a long time. The determination of a sample takes about 1.5 h
The spectrophotometric determination of methionine content is pursued by everyone because of its simple operation, but most of the spectrophotometric methods use the ninhydrin method, and other amino acids have similar reactions, so it will interfere with the determination of methionine content

Method used

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  • Fast plate method for screening of methionine producing bacteria
  • Fast plate method for screening of methionine producing bacteria

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Embodiment Construction

[0014] Seed medium: glucose 50 g peptone 30 g yeast extract 5 g urea 3 g KH 2 PO 4 1.5 gK 2 HPO 4 0.5 g MgSO 4 ·7H 2 O 0.5 g H 2 O 1 L pH 7.0-7.2.

[0015] Fermentation medium: glucose 110 g urea 8 g KH 2 PO 4 1 g MgSO 4 ·7H 2 O 0.2 g FeSO 4 ·7H 2 O 0.2 g MnSO 4 ·H 2 O 0.2 g (NH 4 ) 2 SO 4 2 g Biotin 5 μg / 100mL Thiamine 200 μg / 100mL (dispersion) Agar powder 20 g H 2 O 1 L pH 6.5.

[0016] Take 100 μL of the mutagenized bacterial suspension, spread it on a plate containing fermentation medium, and culture at 30 °C for 1-2 d. When the strains grow obviously on the plate medium, inoculate the colonies one by one into two fermentation medium plates in parallel with sterile toothpicks, and mark them well. After culturing at 30 °C for 4 days, 10 μL of 10 % NaOH was added dropwise to each colony, 1 μL of 10 % sodium nitroprusside was added dropwise, and allowed to stand for 20 min, 10 μL of concentrated phosphoric acid was added dropwise, and the reaction was c...

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Abstract

The invention relates to a method for screening of methionine producing bacteria. A method in the prior art usually comprises the steps of transferring single colonies appearing on a flat into a shake flask, performing culturing for some time, and analyzing whether methionine is contained or not in fermentation liquid by methods of a spectroscopical photometric method, a chromatography or amino acid analyzer and the like. The method in the prior art is time-consuming. According to the method provided by the invention, the technological processes are simplified; after the colonies grow in plates and are cultured for some time, a series of sodium nitroprusside reagents are directly added to the surfaces of the colonies according to quantity, and after a reaction, whether the color is red is observed, so as to determine whether the colonies can produce methionine. According to the method disclosed by the invention, the screening time for methionine producing bacteria is shortened, and the screening efficiency is improved.

Description

technical field [0001] The invention relates to the rapid screening and identification of a methionine-producing bacteria. agricultural science and technology. Background technique [0002] Methionine is a sulfur-containing amino acid and is an essential amino acid for humans and livestock. Methionine provides a source of sulfur for humans or livestock during metabolism. Lack of methionine can cause sub-health. The market will use the metabolism of taurine, glutathione and other substances to provide the required sulfur-containing amino acids. [0003] The source of methionine is mainly chemical synthesis in the current market. Fermentative synthesis of methionine is favored because it can produce pure L-methionine, reduce costs, and reduce pollution. [0004] In order to obtain methionine-producing strains, the commonly used screening method is as follows: obtaining a single colony by streaking on a plate, inoculating the single colony into a liquid medium, fermenting f...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/04
CPCC12Q1/04
Inventor 胡翠英李良智邱业先王桃云钱玮陈宏伟扶教龙
Owner 南京先华生物科技有限公司
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