Method for improving screening efficiency of curdlan gum high-yielding strains

A high-yield strain and screening efficiency technology, which is applied in the direction of bacteria and mutant preparation, can solve the problems of restriction and screening of high-yield strains, shorten the screening cycle, improve the efficiency of plate screening, and improve the blue color of the plate background Effect

Active Publication Date: 2019-07-12
EAST CHINA NORMAL UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The plate screening of mutant strains is a very important link in the mutagenesis breeding process. The traditional plate screening is blind and random, which has great obstacles and limitations to the screening of high-yield strains. Therefore, an effective plate screening method should be established to improve Bacteria screening efficiency and shortening the screening cycle are very important

Method used

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  • Method for improving screening efficiency of curdlan gum high-yielding strains
  • Method for improving screening efficiency of curdlan gum high-yielding strains
  • Method for improving screening efficiency of curdlan gum high-yielding strains

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Embodiment 1 plate selection medium NaI and NaIO 3 determination of concentration

[0051] According to different drug ratios, NaI and NaIO 3 Add the aniline blue screening medium whose pH is 6.0, and the screening medium composition is: ethylene glycol 7%, yeast powder 0.2%, aniline blue 0.005%; NaI and NaIO 3 Concentration (mol / L) ratio: 0.05 / 0.01, 0.075 / 0.015, 0.1 / 0.02, 0.15 / 0.03, 0.2 / 0.04, the pH of 2% agar was adjusted to 7.0; The growth of the colony on the plate under the above conditions and the plate without adding drugs is as follows: figure 1 As shown, the statistical situation of the number of colonies on the different condition plates is shown in Table 1. The number of plate colonies in the control group is more than that of the experimental group, indicating that the plate of the experimental group has a certain lethal effect; the plate colonies of the control group not only turn blue, but also turn blue in large areas, while the plate colonies of the e...

Embodiment 2

[0055] NaI and NaIO when embodiment 2NTG concentration is 0.3mg / mL 3 Influence on screening results of mutagenic strains

[0056] Use 20mM Tris maleic acid buffer with a pH value of 8.5 to prepare 0.3mg / mL NTG for chemical mutagenesis. The plate screening medium is: ethylene glycol 7%, yeast powder 0.2%, aniline blue 0.005%, added NaI with NaIO 3 Concentration ratio is 0.1 / 0.02mol / L, agar 2%, pH is adjusted to 7.0, and mutagenic bacteria liquid is coated control group plate (without adding medicine) and experiment group plate (adding medicine) simultaneously, control group and experiment group respectively pick 50 strains of bacteria, according to the output of the crude extraction of the selected strains in the later stage of fermentation, 16 positive mutant strains in the control group were calculated, with a positive mutation rate of 31%; 24 positive mutant strains in the experimental group, with a positive mutation rate of 47%. The positive mutation rate of the experimen...

Embodiment 3

[0058] NaI and NaIO when embodiment 3NTG concentration is 0.5mg / mL 3 Influence on screening results of mutagenic strains

[0059] The concentration of NTG was 0.5mg / mL, the pH of the plate screening medium was 7.0, and the added NaI and NaIO 3 The concentration ratio is 0.1 / 0.02mol / L, and the mutagenizing bacteria liquid is coated on the control plate (without drug addition) and the experimental group plate (with drug addition) at the same time, 50 strains of bacteria are picked in each of the control group and the experimental group, and the positive mutant bacteria in the control group are There were 14 strains with a positive mutation rate of 28%; in the experimental group, there were 23 positive mutant strains with a positive mutation rate of 46%.

[0060] The statistical situation of the number of colonies of the mutagenized strains on the above-mentioned condition plate is shown in Table 3, and the crude extraction yield of the bacterial strains selected by the control ...

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Abstract

The invention relates to a method for improving the screening efficiency of curdlan gum high-yielding strains, and belongs to the microbial breeding technical field, wherein the method mainly includesthe following steps: (1) culturing starting curdlan gum-producing strains; (2) carrying out NTG mutagenesis of the starting strains; (3) preparing an aniline blue flat plate screening culture medium;(4) adding NaI and NaIO3 into the aniline blue flat plate screening culture medium; (5) carrying out initial screening of a mutagenic bacterial liquid on the aniline blue and NaI and NaIO3 double-screening flat plate culture medium; (6) carrying out fermentation re-screening with a 50 mL / 250 mL shaking flask, and screening high-yielding strains according to the measured crude yield; and (7) carrying out fermentation secondary re-screening with a 75 mL / 500 mL shaking flask, and screening the high-yielding strains according to the measured yield of curdlan gum. The chemical drugs NaI and NaIO3are added to the flat plate screening culture medium, the principle of producing elemental iodine in an acidic environment to kill acid-producing bacteria is used, the probability of screening the curdlan gum high-yielding strains is increased, the efficiency of the flat plate screening bacteria is improved, and the period of screening bacteria is shortened.

Description

technical field [0001] The invention relates to the technical field of microbial breeding, in particular to a method capable of improving the screening efficiency of curdlan high-yield strains. Background technique [0002] Curdlan gum (Curdlan) is an exopolysaccharide produced by microbial fermentation. It has the unique properties of plant polysaccharides and synthetic polymer substances and is safe and non-toxic. After decades of research and development, it has been used in many industrial field. Curdlan gum has a huge market demand at home and abroad, and the application demand in food, chemical, health care, medical and other fields is increasing day by day. The main market share is occupied by the United States and Japan, while the research level of curdlan gum in my country is mainly in the laboratory In the research stage, although factories have started large-scale industrial production, there is still a certain gap between its output and quality compared with fore...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/01C12N1/20
CPCC12N15/01C12N1/20
Inventor 陆泽赟马红豆高红亮江文正常忠义金明飞黄静步国建
Owner EAST CHINA NORMAL UNIV
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