SU1472493A1 - Strain of providencia stuartii bacteria as producer of restrictase - Google Patents

Abstract

The invention relates to the microbiological industry and is a strain that is used to produce PST restrictases 1. The purpose of the invention is to obtain a new strain having a higher level of restrictase synthesis. The proposed strain PROVIDENCIA STUARII 17 VKPM B-4076 yields restriction enzyme PST of 1 4000-45000 u / g biomass or 300000-400000 u / l of culture fluid and has high stability, namely, it retains the ability to produce the enzyme during storage for at least 5 years without selection work. As a result of the use of the proposed invention, the yield of restriction enzyme PST 1 increases from 400-600 U / g biomass to 40000-45000 U / g biomass or from 3000-4800 U / l of culture liquid to 300000-400000 U / l of culture liquid. 1 tab.

Description

not formed. Tests for methyl red and feiylalanine deaminase are positive, and ornithine decarboxylase and Voges-Proskauer reactions are negative. Indole is formed, and hydrogen sulfide is not formed. Strict aerobes. Cultural features. The strain grows on solid nutrient medium containing 3.5% dry nutrient agar and water at 35-37 C for 16-18 h.

The growth of the Providencia stuartii (17) VKPM B-4076 strain is moderate, the stroke is chetiform. The nature of the edge of the colonies is smooth, the gloss of the surface of the colonies is matte, the surface character is smooth.

The profile of the stroke is convex. Optically, the colonies are opaque, i.e. do not project

The strain Providencia stuartii 17 has a specific zapdh, the colonies of this strain are oily in consistency.

It grows well on a liquid nutrient medium containing fish hydral for 50 g / l; water up to 1.0 l, pH 7.2 - 7.4. On such a nutrient medium, the strain grows at 35-37 C for 5-6 h.

At the start of 1 gram on a nutrient medium of a given composition the turbidity of the medium occurs, which indicates a strong growth of the culture. Growth is deep and film does not form on the surface. The sediment, consisting of Providencia stuartii cells 17 tons of gougey, when exposed to lysozyme, cell lysis quickly occurs. The amount of sediment is abundant, the color of the precipitate is light gray.

In contrast to the known, the proposed strain yields restriction enzyme Pst of 1 40000-45000 u / g biomass or 300000-400000 u / l of culture; liquids; It has a high stability, namely: it retains the ability to reproduce the enzyme during storage for at least 5 years without selection work. When cultivating a culture on a nutrient medium of known composition pH 7.4; temperature 37 ° С; aeration of 40 L / min. fermentation is carried out until a pH of 8.0 is reached. Biomass yield: 7.5-8.0 g / l of culture fluid. The output of restrictase Pst 1:

20000-22500 u / g of biomass; 150000 - 180000 u / l of culture fluid. A comparison of the stability of the Pst 1 restrictase content during storage is presented in the table.

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five

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five

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Example G. To obtain

A strain with a high content of restriction enzymes Pst G was carried out continuous selection work with the selection of variants that have high productivity of restriction enzyme Pst 1.

Selection was carried out in Petri dishes on solid medium containing 3.5% nutrient cyxofo agar and water.

The verification of the productivity of the Providencia stuartii 17 VKPM B-4076 strain, half a selection, was carried out in comparison with the Providencia stuartii (164) VKPM B-1789 strain. which has been tested under production conditions.

Prepare the medium of the following composition, g / l:

Fish hydrolyzate 50

Water1, 0

The recovery of the Providencia stuartii VKPM B-4076 strain was carried out on dry nutrient agar (dry nutrient agar 35 g / l, water 1, 0 l) in Petri dishes. Incubated in a heat chamber at 37 ° C for 18 hours.

From the surface of the agar in Petri dishes, the culture was transferred with a bacteriological loop to the shaker flasks with a nutrient medium of a given composition, pH 7.4.

Grow seed at 37 ° C for 18 h on a rocking chair.

Fermentation of the culture is carried out on a nutrient medium of this composition at pH 7.4; temperature 37 С; stirring 200 rpm; air supply - 2 air volumes per 1: volume of the nutrient medium, grown to reach 1472493о

Bite-B-4076 optical density compared to the known spore suspension 1.5 opt. units (FEK, a restriction enzyme yield Pst 1, the filter No. 6, a cuvette 0.5 cm). Grows from 400-600 u / g of biomass to

Biomass yield: 7.5-8.0 g / l cul-40000-A5000 u / g. Tural fluid.

Claims (1)

Pst I 40,000 restriction enzyme yield - Invention formula 45,000 u / g biomass; 300,000 400000 units / l of culture fluid. Strain of bacteria Providencia stuarSo, when using ™ B-4076 - the producer of the restriction strain Providencia stuartii VKPMtaz Pst 1.