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A microbial intelligent cascade heat resistance regulation method

A technology of microorganisms and microorganisms, applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve the problems of high energy consumption in the production process, achieve the effects of improving production efficiency, reducing energy consumption, and increasing fermentation temperature

Active Publication Date: 2018-02-27
BEIJING INSTITUTE OF TECHNOLOGYGY
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The purpose of the present invention is to solve the problem of high energy consumption in the production process due to the control of normal temperature fermentation in the industrial microorganism culture process

Method used

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  • A microbial intelligent cascade heat resistance regulation method
  • A microbial intelligent cascade heat resistance regulation method
  • A microbial intelligent cascade heat resistance regulation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1: Design and Construction of Artificial RNA Thermosensitive Switch

[0024] Drawing on the basic structure of natural RNA thermosensitive switch, the structure of artificial RNA thermosensitive switch is mainly composed of RBS sequence (5'-AAGGAG-3'), ARBS (anti-RBS, antisense RBS sequence) and the loop between them. Structural composition. The secondary structure and free energy of RNA thermosensitive switch can be predicted and calculated by The mfold Web Server. The free energy of the RNA thermosensitive switch can be adjusted by changing the matching degree of the stem-loop base sequence, the G-C content and the size of the loop structure, so as to obtain the RNA thermosensitive switch responding to different temperatures. Therefore, according to the free energy simulated by The mfold Web Server, we designed a series of RNA temperature-sensitive switches with different gradient free energies. A library of RNA temperature-sensitive switch elements respondi...

Embodiment 2

[0034] Example 2: Assembly of cascade heat-resistant system

[0035] (1) Assemble a single-function temperature-sensitive heat-resistant system with a single functional heat-resistant element and a specific temperature-sensitive RNA temperature-sensitive switch: promoter-RNA temperature-sensitive switch-heat-resistant functional element, which is suitable for improving the heat resistance of Escherichia coli at a specific high temperature (2) Rational assembly of RNA temperature-sensitive switches with different temperature responses and heat-resistant functional elements with optimal temperature functions: promoter-RNA temperature Sensitive switch (T1°C)-heat-resistant functional element (T1°C)-promoter-RNA thermosensitive switch (T2°C)-heat-resistant functional element (T3°C)-...-promoter-RNA thermosensitive switch (Tn°C )-heat-resistant functional element (Tn°C) (T1°C<T2°C<...<Tn°C), suitable for expressing different heat-resistant functional elements at different temperatu...

Embodiment 3

[0036] Embodiment 3: Construction and assembly of population quantity system

[0037]In this method, a temperature-sensitive population quantity regulation system is constructed by integrating a quorum sensing system and an RNA temperature-sensitive switch. The RNA thermometer (RNAT12) and the promoter pluxR, which senses AHL concentration, constitute an AND-gate switch that responds to density and temperature. Microorganisms can grow freely during the fermentation process and are no longer limited by the concentration of quantity regulation. However, when the density is high enough, the heat released by microorganisms will increase accordingly. When the metabolic heat causes the system temperature to reach a certain value, the RNA thermometer will be turned on. The expression of the downstream toxin protein is initiated, resulting in cell death, but the cell concentration is not unlimited, and the final cell concentration will be controlled at the cell concentration set by th...

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Abstract

During fermentation, microbial growth and production are often limited by heat stress. For the purpose of constructing a heat-resistant Escherichia coli chassis host, this patent uses synthetic biology methods to create a microbial intelligent cascade heat-resistant regulation method, which can improve the heat resistance of microorganisms at the two levels of cells and flora, which mainly includes Cascade heat-resistant system and quantity control system. The cascade thermotolerance system includes heat-shock proteins (HSPs), ubiquitin and transcription factors and other heat-tolerance functional elements. The combination of artificially designed RNA temperature-sensitive switch and promoter is used as the regulatory element to realize the cascade thermotolerance of microorganisms. The microbial population control system uses the combination of quorum sensing system and RNA temperature-sensitive switch as an AND gate switch to dynamically open the altruistic programmed death system, effectively control metabolic heat, and improve the ability of microorganisms to adapt to heat stress at the level of flora. Through the synergistic effect of the two systems, the adaptability and robustness of microorganisms to specific high temperatures and high temperature fluctuations are improved.

Description

technical field [0001] The invention relates to improving the heat resistance of Escherichia coli and its industrial application by using a constructed microbial intelligent cascade heat resistance control method and belongs to the field of biochemical industry. Background technique [0002] In the past 30 years, the attempt of microbial transformation technology in the field of organic chemical synthesis has not only led to extensive theoretical research, but also made great progress in practical applications. Many pharmaceuticals, food additives, vitamins, cosmetics and other fine chemical products with complex chemical synthesis processes can be obtained by microbial transformation technology. Despite numerous citations, microbial transformation also faces a series of potential unresolved problems. One of the serious problems is that microorganisms are often affected by adverse environments during the growth and production process, such as strong acid, strong alkali, hig...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/31C12N15/113C12N15/09C12R1/19
Inventor 李春贾海洋孙欢孙翔英冯旭东
Owner BEIJING INSTITUTE OF TECHNOLOGYGY
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