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Method for screening and culturing Rahnella sp.R3 producing low-temperature lactase

A Lahnella and screening method technology, applied in the direction of microorganism-based methods, biochemical equipment and methods, enzymes, etc., can solve the problem of low catalytic activity and achieve the effect of optimizing fermentation conditions

Inactive Publication Date: 2013-03-13
JIANGNAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Most of the commercialized lactases are normal temperature type, the optimum reaction temperature of the enzyme is 37°C or higher, and the catalytic activity is very low at low temperature

Method used

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  • Method for screening and culturing Rahnella sp.R3 producing low-temperature lactase
  • Method for screening and culturing Rahnella sp.R3 producing low-temperature lactase

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1: Screening method for Rahnella sp.R3 producing low-temperature lactase

[0022] (1) Primary screening: absorb the enriched culture solution and transfer it to the primary screening plate at different dilutions by the doubling dilution method, spread evenly, culture at 15°C, screen blue colonies, separate and purify, and store on the slant.

[0023] (2) Re-screening: select the upper limit temperature as 40°C, that is, strains that do not grow at this temperature are low-temperature bacteria, and select this bacteria as the test strain. The test strains preserved on the slant were activated, and the transfer fermentation culture was carried out at 15°C for shake flask culture (50mL / 250mL) for 48h. Extract the crude enzyme solution, measure the hydrolysis activity at 15°C, and select the bacteria R3 with higher activity as the test strain.

Embodiment 2

[0024] Embodiment two: Rahnella R3 (Rahnella sp.R3) strain characteristics

[0025] (1) Growth characteristics: the optimum growth temperature of the strain is 15°C; the optimum growth pH is 7

[0026] (2) Bacterial characteristics: Gram-negative, short bacilli, 1.5 μm to 2 μm in length, capsulated, without spores.

[0027] (3) Colony characteristics: The colony is yellow, round and raised, sticky in texture, smooth in edge, opaque, and easy to pick.

[0028] (4) Physiological and biochemical characteristics: fermenting D-glucose to produce acid and gas, gelatin hydrolysis test was negative, lysine and ornithine decarboxylase and arginine dihydrolase were negative, nitrate reduction, MR (methyl Red), VP (Volts-Prauter's test) were positive. Fermentable lactose, maltose, sucrose, L-rhamnose, raffinose, cellulose sugar.

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Abstract

The invention relates to a screening and culturing method of Rahnella sp.R3. The method adopts a bacterial enrichment medium (1 / 4TSB) and lactose as substrates, takes X-Gal as a color developing agent, and performs agar identification plate primary screening and fermentation enzymatic activity determination secondary screening to obtain the high-yield low-temperature lactase strain Rahnella sp.R3. Single factor experiment and orthogonal experiment analysis is conducted to obtain the optimal fermentation enzyme production formula and fermentation condition, and finally the flask-shaking fermentation level can reach 5.92U / mL.

Description

Technical field [0001] The invention relates to a Rahnella sp. R3 isolated from permafrost in Tianshan, Xinjiang; the invention relates to a method for screening and culturing the Rahnella sp. R3. Background technique [0002] Lactase (β-D-galactoside hydrolase, EC3.2.1.23), which can be derived from plants, animals and microorganisms, can catalyze the hydrolysis of β-galactosidic bonds in lactose into glucose and galactose. Lactose is the most important carbohydrate in milk, with a content of about 4.5% to 5.0%. Some people cannot absorb and utilize lactose, which can cause lactose intolerance. At present, the best way to solve lactose intolerance is to use lactase to hydrolyze lactose to produce low-lactose or lactose-free dairy products. Most of the commercialized lactases are normal temperature type, the optimum reaction temperature of the enzyme is 37°C or higher, and the catalytic activity is very low at low temperature. Since most operations of dairy processing are...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/04C12N9/38C12N1/20C12R1/01
Inventor 杨瑞金沈莲莲华霄
Owner JIANGNAN UNIV
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