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Method for high-throughput screening of high-yield sophorolipid strains

A sophorolipid, high-throughput technology, applied in the field of high-throughput screening of high-yielding sophorolipid strains, can solve the problems of single screening pressure, easy degradation, slow growth of strains, etc., to increase mutation rate and positive mutation rate, increase Mutation rate and positive mutation rate, the effect of increasing the positive mutation rate

Pending Publication Date: 2020-08-21
EAST CHINA UNIV OF SCI & TECH +1
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the characteristics of sophorolipid itself, high oxygen consumption and high viscosity in the fermentation production process lead to increased production costs. Therefore, it is necessary to screen out high-yield sophorolipid production strains
[0003] Rational genetic engineering breeding method is a favorable method for strain improvement, but it mainly has the following disadvantages: (1) cell metabolism is a very complex adaptive network, changing one or two genes may not be able to achieve the required goal; (2) ) The production and sales of some genetically engineered strain-related products are restricted by relevant application fields
In the traditional high-throughput screening method, such as the research results of Zhou Gang et al., it can be seen that the strains obtained by the traditional mutagenesis method grow slowly and easily degenerate, and the screening efficiency is not high; Zhou Gang et al. Only a single screening pressure was used in the process, and the resulting high-yielding strains were more likely to degrade during the actual fermentation process.

Method used

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  • Method for high-throughput screening of high-yield sophorolipid strains
  • Method for high-throughput screening of high-yield sophorolipid strains
  • Method for high-throughput screening of high-yield sophorolipid strains

Examples

Experimental program
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Embodiment 1

[0057] The present embodiment provides a method for high-throughput screening of high-yield sophorolipid strains, the method comprising:

[0058] The step of strain activation;

[0059] Atmospheric room temperature plasma (ARTP) combined mutagenesis steps with sodium nitrite;

[0060] Steps for relaxation culture;

[0061] Steps of multiple stress culture;

[0062] Wherein, the bacterial species is Candida; in the step of multiple pressure culture, the combination of iodoacetic acid and malonic acid is used to carry out double pressure culture screening.

[0063] In this embodiment, the step of activating the strains includes: taking the strains in a glycerol tube and connecting them to the activation medium, culturing them in shake flasks at 220r / min and 25°C for 48 hours to activate the strains, and diluting the strains to a concentration of 10 7 cells / ml to obtain the activated bacterial liquid.

[0064] In this embodiment, the step of combined mutagenesis of atmospheri...

Embodiment 2

[0069] The present embodiment provides a method for high-throughput screening of high-yield sophorolipid strains, the method comprising:

[0070] The step of strain activation;

[0071] Atmospheric room temperature plasma (ARTP) combined mutagenesis steps with sodium nitrite;

[0072] Steps for relaxation culture;

[0073] Steps of multiple stress culture;

[0074] Wherein, the bacterial species is Candida; in the step of multiple pressure culture, the combination of iodoacetic acid and malonic acid is used to carry out double pressure culture screening.

[0075] In this embodiment, the step of activating the strains includes: taking 200 μl of the glycerol tube strains and connecting them to the activation medium, placing them in a 500 ml shake flask, and culturing the strains at 220 r / min and 25° C. for 48 hours to activate the strains, Dilute to a concentration of 10 7 cells / ml to obtain the activated bacterial liquid.

[0076] In this embodiment, the step of combined m...

Embodiment 3

[0087] Comparing Example 3 with Example 2, the only difference is that the combined mutagenesis conditions are 10 min, 5 mg / L sodium nitrite mutagenesis conditions combined with ARTP mutagenesis for 18 s.

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Abstract

The invention discloses a method for high-throughput screening of high-yield sophorolipid strains. The method comprises the following steps: activating strains; carrying out combined mutagenesis on normal-pressure room-temperature plasma and sodium nitrite; carrying out relaxation culture; and carrying out multi-pressure culture; wherein the strain is candida mycoderma bacteria; in the step of multi-pressure culture, iodoacetic acid and malonic acid are combined for dual-pressure culture screening. According to the method, normal-pressure room-temperature plasma and sodium nitrite are combinedfor mutagenesis, relaxation culture is combined with double pressures of iodoacetic acid and malonic acid for screening, the mutation rate and the positive mutation rate of the strain are increased,the probability of obtaining the high-yield strain is increased, and then the high-yield strain with good genetic stability is obtained.

Description

technical field [0001] The invention relates to the technical field of microbial fermentation, in particular to a method for high-throughput screening of high-yield sophorolipid strains. Background technique [0002] Sophorolipid is a kind of glycolipid biosurfactant. Although there are many kinds of microorganisms that can synthesize sophorolipid, Candida bombicola is the main producer in the research report. Sophorolipids have attracted much attention in industrial production and scientific research, and are considered to be one of the most promising bioactive agents due to their environmental friendliness. Due to the characteristics of sophorolipid itself, high oxygen consumption and high viscosity in the fermentation production process lead to increased production costs. Therefore, it is necessary to screen out high-yield sophorolipid production strains. [0003] Rational genetic engineering breeding method is a favorable method for strain improvement, but it mainly has...

Claims

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Application Information

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IPC IPC(8): C12N15/01C12N1/20C12N1/36C12R1/72
CPCC12N15/01C12N1/20C12N1/36
Inventor 储炬李前会田锡炜杭海峰夏建业庄英萍李璟曦张嘉兴韩思宇徐文静杨露露
Owner EAST CHINA UNIV OF SCI & TECH
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