Recombined corynebacterium glutamicum as well as preparation method and application thereof

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

Active Publication Date: 2018-05-29
JIANGNAN UNIV
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  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

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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  • Recombined corynebacterium glutamicum as well as preparation method and application thereof
  • Recombined corynebacterium glutamicum as well as preparation method and application thereof
  • Recombined corynebacterium glutamicum as well as preparation method and application thereof

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Embodiment 1

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

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

[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 on sgRN...

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Abstract

The invention provides recombined corynebacterium glutamicum, can be used for effectively solving the technical problem that the conventional corynebacterium glutamicum is low in foreign protein expression quantity. The peptidoglycan endonucleases gene mepA, cytomembrane surface anion protein gene porB and endoproteinase gene clpC in the corynebacterium glutamicum are knocked out, meanwhile ACB transporter gene Ncg10909 are over expressed. Moreover the invention further provides a preparation method for the recombined corynebacterium glutamicum and application thereof.

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 Applications(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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