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Method for screening bacterial strains with high output of low-temperature MDH (malate dehydrogenase)

A technology of malate dehydrogenase and bacterial strains, applied in the field of fermentation engineering, to achieve the effect of narrowing the scope

Inactive Publication Date: 2018-12-11
DALIAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] Since there is no specific screening method for the screening of strains producing MDHs at present, it is necessary to study its screening method for the research of high-production MDHs

Method used

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  • Method for screening bacterial strains with high output of low-temperature MDH (malate dehydrogenase)
  • Method for screening bacterial strains with high output of low-temperature MDH (malate dehydrogenase)
  • Method for screening bacterial strains with high output of low-temperature MDH (malate dehydrogenase)

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Embodiment 1

[0026] The present invention screens the method for high-yielding low-temperature malate dehydrogenase bacterial strain, specifically as follows:

[0027] (1) Preliminary screening of strains: Take sea mud and seawater samples near the Bohai Sea area of ​​Dalian, Liaoning, China, dilute and smear the plate in beef extract peptone solid medium containing 0.1g / L bromocresol green, and culture at a constant temperature at 25°C 3-4 days; according to the yellowish color of the yellow-green circle generated around the colony and the ratio of the diameter of the color circle to determine the primary screening high-activity target strain, use the method of three rounds of streaking to inoculate on beef extract peptone solid medium ( Beef extract 3g / L, peptone 10g / L, agar 20g / L, seawater configuration, pH 7.2) for purification culture, 25°C, constant temperature culture for 1-2 days, repeat this step 2-4 times;

[0028] (2) Re-screening of strains: Dip the above-mentioned colonies wit...

Embodiment 2

[0031] The present invention screens the method for high-yielding low-temperature malate dehydrogenase bacterial strain, specifically as follows:

[0032] (1) Preliminary screening of strains: Take sea mud and seawater samples near the Yellow Sea area of ​​Qingdao, Shandong, China, dilute them in beef extract peptone solid medium containing 0.15g / L bromocresol green, and culture at a constant temperature at 25°C 3-4 days; according to the yellowish color of the yellow-green circle formed around the colony and the ratio of the diameter of the color circle to judge the target strain with high activity in the primary screening, use the method of three rounds of streaking to inoculate on the beef extract peptone solid medium Carry out purification culture, 25 ° C, constant temperature culture for 1-2 days, this step is repeated 2-4 times;

[0033] (2) Re-screening of strains: Dip the above-mentioned colonies with an inoculation needle to pick up a single colony, puncture and inocu...

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Abstract

The invention relates to the field of fermenting engineering, in particular to a method for screening bacterial strains with high output of low-temperature MDH (malate dehydrogenase). The MDH is a keyenzyme for cell center oxidizing channels, and is closely related with the movement of bacterial flagellum in related reports. The method has the advantages that the bromocresol green is used for primary screening, and the bacterial strain with stronger acid production property is screened out; the power property of the bacterial strain is detected by MTT (methyl thiazolyl tetrazolium), the screening is performed for the second time, and the bacterial strain with stronger power property is selected to test the activity of the enzyme; the low-temperature bacterial strain with high output of MDHs can be accurately screened out by double screening; finally, the activity of the enzyme is detected to finally determine the high-output bacterial strain; the bacterial strains with high output oflow-temperature intracellular MDH can be conveniently and accurately screened out within 20 days, and the blank in the related field of screening of culture mediums by the MDH is filled.

Description

technical field [0001] The invention relates to the field of fermentation engineering, in particular to a method for screening high-yielding low-temperature malate dehydrogenase bacterial strains. Background technique [0002] MDHs (low temperature malate dehydrogenase), as a key enzyme in the central metabolic pathway of organisms, has important research value in genetic variation and individual development, and is widely used in clinical diagnosis and industrial testing. The market demand is increasing day by day, and the value is huge . Since the 1950s, some scholars have isolated MDHs from animal myocardium. At present, the sources of commercial MDHs on the market are mainly extracted from the myocardium, liver, and skeletal muscle of pigs, rabbits, and cattle. The cost of raw materials is low and the source is wide, but the extraction process is cumbersome and the enzyme activity of the product is low. Therefore, it is particularly important to develop new sources. ...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12R1/07C12R1/43
CPCC12N1/20
Inventor 张庆芳肖景惠王梦雨迟乃玉逄飞
Owner DALIAN UNIVERSITY
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