SU1254006A1 - Strain pseudomonas species m7 as test culture for determining l-thyrosine - Google Patents

Abstract

The strain Pseudomonas species M7 (collections of the Central Museum of Industrial Microorganisms of the Institute of Scientific Research Institute of Genetics, collection number B-3245) is a test culture for determining L-tyroein. JV

Description

The invention relates to the microbiological industry and concerns a native strain, which is a test culture for determining the production of L-tyrosine.

The aim of the invention is to obtain a new strain of Pseudomonas species M7 that is specific and sensitive to the content of L tyrosine in the medium with which t-tyrosyc can be detected, starting with 0.5 µg / ml, on simple liquid and agarized mineral media. among various groups of microorganisms, including those assimilating lower alcohols (ethanol or methanol) as a source of carbon and energy

The proposed strain Pseudomonas sp. M7 was obtained by assisting mutagenesis. M-methyl-K-nitro-K-nitrosoguanidine is an auxotrophic tyrosine avi mutant of the wild-type strain Pseudotnonas sp. M isolated from the sewage of the Minsk plant of medical preparations.

Identification of tyrosine dependence (TYR) mutations in bacteria Pseudomonas sp. M7 was carried out by the lack of plant ability on mineral media without L-tyrosine. The minimum concentration of L-tyrosine, which ensures the growth of bacteria of this strain in liquid and agarized mineral media, is 0.5 µg / ml.

Strain Pseudomonas sp. M7 is stored in the Central Museum of Industrial Microorganisms VNGgenetics (Moscow) No. В-3245.

Strain Pseudornonas sp. M 7 has the following characteristics.

.Morphological features of the strain. Cells are straight, rod-shaped with rounded ends 0.6 x 2-3 p. motile with 2-4 monopolar flagella, gram-negative, do not form a dispute.

Cultural features. Colonies on agar nutrient media (co-peptone agar, King In agar) are smooth, flat, round, the edge is flat, grayish-white. After 24 hours at 28 ° C, colonies measuring 2–3 mm are formed. The growth of the injection in Ш1А is moderate, mainly on the surface. Wednesday :. In liquid nutrient media forms intense turbidity.

It grows well on mineral media of simple composition, supplemented with L tyrosine, and does not require the addition of growth factors and other amino acids.

Physiological and bkohimicheskie Signs.

In relation to oxygen - obligate aerobic. The optimum growth temperature is 28-30 ° C, the pH of the medium is in the range of 6.8-7.3. Nitrogen is utilized in the form of ammonium, urea, and nitrate salts.

Pigments. Forms a yellow-green water-soluble fluorescent pigment on King In agar. Intracellular pigment does not form.

The reaction to oxidase and catalase is positive. The reaction of Fores Proskauer negative. Indole does not form. The test for anaerobic arginine-dehydrol is positive. The reaction to anaerobic denitrification is negative. Leo citinase activity is absent.

Does not hydrolyze starch, gelatin, tween-80.

As carbon sources uses glucose, glycerin, sucrose,

5 galactose, maltose, fructose, ethanol, methanol, tyrosine, phenylalanine, ornitin, glycine, leutzn, glutamic acid, aspartic acid. Does not utilize arabinose, mannose, xylo. bite, lactose, mannitol, raffinose, dulcite, sorbitol, cellobiose, inositol, serine, lysine, tryptophan, anthranilic acid, acetamide.

The strain is identified as Pseudomonas sp, M7 according to the Bergey determinant.

Example 1. Determination of L-tyrosine using the Pseudomonas sp. M7 is produced as follows. Do seeding culture Pseudomonas sp. M7 from scythe in MPB (m soy peptone broth), or in another nutrient medium providing the growth of bacteria, and cultured for

, nights at 28 ° C. Bacterial cells are then removed twice with physiological saline by centrifugation (10 minutes, 5000 g) and resuspended to their original volume in the same solution. 0.1 ml of cell suspension are introduced into a test tube with 4 ml of minimal semi-liquid agar (0.7%) melted and cooled to 45 ° C, prepared on the basis of M9 mineral medium containing glucose as a carbon source (at a concentration of 0.2 %); and layering with the second layer on the surface of the agar medium (1.5%) of the same composition.

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After the inoculation of the upper layer with the inoculation culture, individual cultures of the clones tested for the production of L-tyrosine are cultured by microorganisms that utilize glucose as a source of carbon and energy (20-30 clones per Petri dish). Samples are grown for 24 - 48 h at 8-30 ° C. Clear, intense growth zones of the Pseudomonas sp, M7 indicator test culture are observed around the colonies whose cells produce L-tyrosine, the size of which depends on the amount of L-thyroid consciousness synthesized.

Example 2. Sowing material of the test culture of Pseudomonas sp. M7 and the following procedures are produced as in Example 1, except that ethanol (at a concentration of 1%) is added to the mineral media used instead of glucose as the carbon source and energy, and the test samples are cultured for L-tyrosine cultures. microorganisms that recycle ethanol.

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PRI me R 3. Sowing material test culture Pseudoraonas sp. M7 and the following procedures are produced as in Example 1, except that methanol (at a concentration of 1%) is added to the used minerals as a source of carbon and energy, and cultures of microorganisms that utilize methanol are seeded for L-tyrosine. Samples are grown for 48-72 h.

Strain Pseudomonas sp. M7 is a mutant for the TYR gene, which ensures its need and high specificity for L-tyrosine. Using strain Pseudomonas sp. M7 can be used to view and study the production of L-tyrosine, starting with 0.5 µg / ml, on simple mineral media among the various groups of MiiKpoopraznam, including assimilating lower alcohols (ethanol or methanol) as carbon sources and energy.

Claims (1)

Example Seed material test-culture RzeiPotpopav sp. M7 and the subsequent procedures are carried out as in Example 1, except that methanol (at a concentration of 1Ζ) is added as a carbon source and energy to the mineral media used, and the cultures of microorganisms that utilize methanol are seeded for the content of L-tyrosine. Samples are grown for 48-72 h. Strain RzeiPotopaz vr. M7 is a mutant for the ΤΥΕ-gene, which ensures its need and high specificity for b-tyrosine. Using strain RweiPotopaz sp. M7 can be used to identify and study the production of L-tyrosine, starting at 0.5 µg / ml, on simple mineral media among various groups of microorganisms, including those assimilating lower alcohols (ethanol or methanol) as sources of carbon and energy.