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Yeast genetically engineered strain expressing hrp gene and constructing method thereof

A technology of genetically engineered strains and construction methods, applied in the field of yeast genetically engineered strains and their construction, can solve the problems of small harvest, complicated process, and inability to meet market needs, etc.

Inactive Publication Date: 2003-11-12
WUCAI FINE CHEM JIANGSU
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] At present, there are two ways to obtain the Harpin protein. One is to extract the protein from plant pathogenic bacterial cultures, but the process is complex, the harvest is small, and the price is high, which is far from meeting the needs of the market; the other is to use genetic engineering. Method, that is, through the fermentation of genetically engineered strains, a large amount of biologically active Harpin protein is produced, and it is used as an active ingredient of plant antibacterial and bacteriostatic agents for crop protection
At present, the production of Harpin protein reported at home and abroad is based on Escherichia coli as the final expression vector, and IPTG is needed as an inducer to promote the expression of the target protein. The inducer is more expensive and the cost of industrial production is higher.
There is no report on the production of Harpin protein using yeast as the final expression vector

Method used

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  • Yeast genetically engineered strain expressing hrp gene and constructing method thereof
  • Yeast genetically engineered strain expressing hrp gene and constructing method thereof
  • Yeast genetically engineered strain expressing hrp gene and constructing method thereof

Examples

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Embodiment 1

[0108] Construction method of genetic engineering strain HrpZ

[0109] 1. Genomic DNA extraction and hrpZ gene isolation and cloning

[0110] 1. Genomic DNA Extraction

[0111] A strain of Pseudomonas syringae was isolated and identified from the soil, and the genomic DNA was extracted according to the instructions of the Genomic DNA Extraction Kit from QIAGEN Company.

[0112] 2. Isolation of hrpZ gene

[0113] The hrpZ gene was amplified and isolated by PCR using the genomic DNA of Pseudomonas syringae as a template, and the PCR primer sequences were as follows:

[0114] Forward: 5'-ATGCAGAGTCTCAGTCTTAACAGCA-3'

[0115] Reverse: 5'-AGTTCTCCGTCAGGCGGTTGTC-3'

[0116] PCR reaction conditions:

[0117] Pre-denaturation: 94°C for 1 minute

[0118] Denaturation: 92°C for 45 seconds

[0119] Refolding: 66°C for 45 seconds

[0120] Extension: 68°C for 2 minutes

[0121] Cycles: 30

[0122] Termination of extension: 68°C for 20 minutes

[0123] The PCR product was detecte...

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Abstract

The present invention relates to a yeast gene engineering strain capable of expressing hrp gene and its construction method. The name of said strain is Saccharomyces cerevisiae hrp Z, its number is CCTCC No:M203004. Its construction method includes: extraction of pseudomonas syringae genome DNA; using genome DNA as template PCR primer to make construction of hrp Z recombinant yeast gene engineering strain; hrpZ gene contained yeast conversion and induction. The gene engineering harpin protein produced by said strain can induce plant hypersensitive reaction, activate internal self-defense function of plant and promote plant growth. so that it can raise yield and quality of crops.

Description

technical field [0001] The invention relates to bioengineering, in particular to a yeast gene engineering strain expressing hrp gene and its construction method. Background technique [0002] The hrp gene cluster exists in a variety of plant pathogenic bacteria, and the Harpin protein encoded by it is a protein necessary for bacterial pathogenicity, but for non-host plants, it can cause hypersensitive responses of plants (Hypersensitive response), its Expressed by rapid, localized atrophy and death of the infected tissue, this limits the possibility of further spread of the pathogen. Hypersensitivity response is a kind of protective response that plants have, which is similar to the broad-spectrum immune response against pathogens and pests in higher animals. The Harpin protein itself has no killing or inhibiting effect on bacteria, viruses, fungi, nematodes, and insects. Instead, it binds to plant receptors and generates a certain signal to stimulate or induce the plant's ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/19C12N1/20C12N5/10C12N15/31C12N15/70C12N15/81C12P21/00
Inventor 魏晨飞
Owner WUCAI FINE CHEM JIANGSU
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