A kind of recombinant Corynebacterium glutamicum, its preparation method and application

A glutamic acid rod and bacillus technology, applied in the biological field, can solve the problem of low expression of exogenous protein, and achieve the effect of increasing the expression

Active Publication Date: 2021-08-24
JIANGNAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] In view of the above problems, the present invention provides a recombinant Corynebacterium glutamicum, which can effectively solve the technical problem of low expression of exogenous proteins of the existing Corynebacterium glutamicum. In addition, this method also provides the recombinant glutamic acid Preparation method and application of coryneform bacteria

Method used

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  • A kind of recombinant Corynebacterium glutamicum, its preparation method and application
  • A kind of recombinant Corynebacterium glutamicum, its preparation method and application
  • A kind of recombinant Corynebacterium glutamicum, its preparation method and application

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Embodiment 1

[0037] 1. Construction of three gene knockout vectors and one gene overexpression vector of Corynebacterium glutamicum

[0038] The nucleotide sequence of porb is shown in Gene ID: 1018962, the nucleotide sequence of mepa is shown in Gene ID: 1020444; the nucleotide sequence of clpc is shown in Gene ID: 1020624; the nucleotide sequence of Ncgl0909 is shown in GeneID: 1018938 .

[0039] Corynebacterium glutamicum C. glutamicum The relevant genes in the ATCC 13032 genome were used as templates to design sgRNA.

[0040] Wherein, the Gene ID of porb nucleotide sequence, mepa nucleotide sequence and clpc nucleotide sequence and C. glutamicum The nucleotide sequences of ATCC 13032 are all published.

[0041] The sgRNA used for knockout should have a 5′-(N 20 )-NGG-3′ structure, and located in the gene sequence, design sgRNA corresponding to mepa, prob, clpc, as shown in the sequences SEQ ID No.1, No.2 and No.3 respectively. Two oligonucleotide strands were synthesized based o...

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Abstract

The invention provides a recombinant coryneform bacterium glutamicum, which can effectively solve the technical problem of low expression of exogenous proteins of the existing coryneform bacterium glutamicum. The endopeptidoglycanase gene mepA, the anion channel protein gene porB on the cell membrane surface and the intracellular protease gene clpC in the recombinant Corynebacterium glutamicum were all knocked out, and the ABC transporter gene Ncgl0909 was overexpressed. In addition, the method also provides the preparation method and application of the recombinant Corynebacterium glutamicum.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a recombinant coryneform bacterium glutamicum, its preparation method and application. Background technique [0002] Corynebacterium glutamicum belongs to the genus Corynebacterium and is a Gram-positive bacterium. It is generally believed that Corynebacterium glutamicum is non-toxic, non-pathogenic, and non-spore-producing, and is generally recognized as a safe strain (GRAS). At the same time, Corynebacterium glutamicum has the advantages of rapid growth, clear genetic background, and good protein secretion system. It has been widely used in the production of amino acids in the past 50 years, such as glutamic acid, lysine, etc., and is also used in food Industry, production of animal feed. In recent years, it has also been used for the expression of medicinal proteins and their precursors. It is a safe host for the expression of medicinal proteins and has high applicat...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/21C12N15/113C12N15/63C12P21/00C12R1/15
CPCC07K14/34C12N9/16C12N9/52C12N15/113C12N2310/10C12P21/00
Inventor 刘秀霞彭枫王新月董贵彬杨艳坤白仲虎
Owner JIANGNAN UNIV
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