Yunnan Tengchong source ectoine synthetic gene cluster and application thereof

A technology for ectoine and transgenic plants, which is applied in the field of high-yield ectoine-producing Escherichia coli engineering bacteria and Paenibacillus ectoine synthesis gene clusters, can solve the problems of limited yield, limited strain synthesis ability, and high production cost. Achieve the effect of enhancing the function of cells, restoring and stabilizing the barrier function of the stratum corneum, and increasing the moisture content of the skin

Active Publication Date: 2021-08-27
深圳中科欣扬生物科技有限公司 +2
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the production of ectoine by these traditional methods is limited by the synthesis ability of the strain itself, and the output is limited. At the same time, the high-salt environment required for production is extremely harmful to the corrosion of production equipment, and the production cost is extremely high, which is not suitable for large-scale industrial production.

Method used

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  • Yunnan Tengchong source ectoine synthetic gene cluster and application thereof
  • Yunnan Tengchong source ectoine synthetic gene cluster and application thereof
  • Yunnan Tengchong source ectoine synthetic gene cluster and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0105] Embodiment 1, the construction of Escherichia coli ectoine high-yielding bacterial strain

[0106] 1. PCR amplification of the coding sequence of genes related to ectoine synthesis

[0107] Using the preserved metagenomic library DNA as a template, PCR amplification was performed with primers F1 and R1 to obtain the PCR amplification product EctA gene, namely fragment 1. The EctA gene contains a DNA molecule whose coding sequence is SEQ ID No.4. The DNA molecule shown in SEQ ID No.4 encodes the EctA protein shown in SEQ ID No.1.

[0108] Using the preserved metagenomic library DNA as a template, PCR amplification was performed with primers F2 and R2 to obtain the PCR amplification product EctB gene, namely fragment 2. The EctB gene contains a DNA molecule whose coding sequence is SEQ ID No.5. The DNA molecule shown in SEQ ID No.5 encodes the EctB protein shown in SEQ ID No.2.

[0109] Using the preserved metagenomic library DNA as a template, PCR amplification was p...

Embodiment 2

[0120] The biotransformation of embodiment 2 escherichia coli ectoine

[0121]1. Shake flask culture

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Abstract

The invention discloses a Yunnan Tengchong source ectoine synthetic gene cluster and application thereof. Specifically, the invention discloses a novel ectoine biosynthetic gene cluster containing encoded aminobutyric acid acetyltransferase (EctA), diaminobutyric acid aminotransferase (EctB) and ectoine synthetase (EctC), and the three genes are integrated into a pBAD / HisA carrier by taking escherichia coli as a chassis cell to construct a genetically engineered bacterium for high yield of ectoine. Gene expression quantity is enhanced by optimizing fermentation conditions, inducer concentration and the like, biotransformation is carried out starting from the raw material sodium aspartate, biosynthesis of ectoine is increased, and the yield of the genetically engineered bacterium for high yield of ectoine can reach 10.02 g / L after the genetically engineered bacterium is cultured for 72 hours in a fermentation tank and reaches the international advanced level.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and relates to a ectoine synthetic gene cluster from Tengchong, Yunnan and its application, in particular to a Paenibacillus ectoine synthetic gene cluster from Tengchong, Yunnan, a recombinant expression vector, and an Escherichia coli project for high-yield ectoine Bacteria and their applications. Background technique [0002] Tetrahydropyrimidine, also known as tetrahydromethyl pyrimidine carboxylic acid (Ectoine, C 6 h 10 N 2 o 2 ), is a heterocyclic amino acid derivative that widely exists in halophilic bacteria and participates in osmotic pressure regulation. As a compatible small molecule, ectoine plays the role of a protector in extreme environments, and plays a positive role in the maintenance of protein activity, nucleic acid conformation, and microenvironmental homeostasis in organisms. As a powerful protective agent, ectoine reduces the damaging effects of external fa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/54C12N15/60C12N9/10C12N9/88C12N15/70C12N1/21C12P17/12C12R1/19
CPCC12N9/1029C12N9/88C12N9/1096C12N15/70C12Y203/01178C12Y402/01108C12Y206/01019C12P17/12C07K2319/00Y02A50/30
Inventor 董亮张山张岩峰魏焘马骁王飞飞苏温柔
Owner 深圳中科欣扬生物科技有限公司
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