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Genetically-engineered high-producing strain streptomyces diastatochromogenes, production method of epsilon-polylysine and application

A technology of Streptomyces chromogenes and polylysine, applied in the direction of genetic engineering, microorganism-based methods, biochemical equipment and methods, etc., can solve the problem of fewer ε-polylysine-producing bacteria and improve the fermentation level Effect

Active Publication Date: 2020-09-04
TIANJIN UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] As mentioned above, although there are reports on the use of genetic engineering technology for the transformation of bacterial strains, there are few reports on ε-polylysine-producing bacteria, and in terms of ε-polylysine and glucose conversion rates, according to Reports from Japan still have a certain distance

Method used

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  • Genetically-engineered high-producing strain streptomyces diastatochromogenes, production method of epsilon-polylysine and application
  • Genetically-engineered high-producing strain streptomyces diastatochromogenes, production method of epsilon-polylysine and application
  • Genetically-engineered high-producing strain streptomyces diastatochromogenes, production method of epsilon-polylysine and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0077] A kind of genetic engineering high-yield amylase strain Streptomyces chromogenes SDHB (Streptomycesdiastatochromogenes SDHB), the construction steps are as follows:

[0078] (1) Obtaining the target fragment gene:

[0079] The primer sequence sdhB-FF / sdhB-RR was designed according to the sdhB gene, and the restriction sites of XbaI and EcoRI were respectively introduced at both ends of the gene sdhB, and 6 nucleotides were added to the upstream 5' end of the sdhB gene nucleotide sequence, Form the site of the restriction endonuclease XbaI, add 6 nucleotides to the downstream 5' end of the sdhB nucleotide sequence to form the site of the restriction endonuclease EcoRI, PCR amplify the amylase Streptomyces chromogenes TUST sdhB gene in;

[0080] The sequence of the primer sdhB-FF / sdhB-RR is:

[0081] sdhB-FF: SEQ No.2, namely 5'- tctaga atggcgcatgtggaccgg-3', the underlined sequence is the XbaI restriction site;

[0082] sdhB-RR: SEQ No.3, namely 5'- gaattc tcaccgg...

Embodiment 2

[0094] A kind of genetic engineering high-yielding amylase strain Streptomyces diastatochromogenes DCDA (Streptomycesdiastatochromogenes DCDA), the construction steps are as follows:

[0095] (1) Obtaining the target fragment gene:

[0096] The primer sequence dcdA-FF / dcdA-RR was designed according to the dcdA gene, and the restriction sites of XbaI and EcoRI were respectively introduced at both ends of the gene dcdA, and 6 nucleotides were added to the upstream 5' end of the dcdA gene nucleotide sequence, Form the site of the restriction endonuclease XbaI, add 6 nucleotides to the downstream 5' end of the dcdA nucleotide sequence to form the site of the restriction endonuclease EcoRI, PCR amplify the amylase Streptomyces chromogenes TUST The dcdA gene in;

[0097] The sequence of the primer dcdA-FF / dcdA-RR is:

[0098] dcdA-FF: SEQNo.5, namely 5'- tctaga gtgcctccgtctgtggagc-3', the underlined sequence is the XbaI restriction site;

[0099] dcdA-RR: SEQNo.6, namely 5'- ...

Embodiment 3

[0111] A kind of genetic engineering high-yield amylase strain Streptomyces chromogenes ASNO (Streptomycesdiastatochromogenes ASNO), the construction steps are as follows:

[0112] (1) Obtaining the target fragment gene:

[0113] The primer sequence asnO-FF / asnO-RR was designed according to the asnO gene, and the restriction sites of XbaI and EcoRI were respectively introduced at both ends of the gene asnO, and 6 nucleotides were added to the upstream 5' end of the asnO gene nucleotide sequence, Form the site of the restriction endonuclease XbaI, add 6 nucleotides to the downstream 5' end of the asnO nucleotide sequence to form the site of the restriction endonuclease EcoRI, PCR amplify the amylase Streptomyces chromogenes TUST asnO gene in;

[0114] The sequence of the primer asnO-FF / asnO-RR is:

[0115] asnO-FF: SEQ No.8, namely 5'- tctaga atgtgcggaatcaccggc-3', the underlined sequence is the XbaI restriction site;

[0116] asnO-RR: SEQ No.9, namely 5'- gaattc tcagagg...

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Abstract

The invention relates to a genetically-engineered strain streptomyces diastatochromogenes with high production of epsilon-polylysine. The streptomyces diastatochromogenes is obtained by over-expressing three key genes, in different metabolic pathways, of a succinate dehydrogenase gene sdhB, a lysine / ornithine decarboxylase gene dcdA and an asparagine synthetase gene asnO in streptomyces diastatochromogenes TUST separately. According to the genetically-engineered strain streptomyces diastatochromogenes with high production of the epsilon-polylysine, by over-expressing the key genes in the different metabolic pathways, the genetically-engineered recombinant strain is obtained, and an experiment proves that the epsilon-polylysine-producing capability of the streptomyces genetically-engineeredstrain is improved compared with that of the original strain streptomyces diastatochromogenes TUST in the same situation, so that the excellent strain is provided for production of epsilon-polylysine.

Description

technical field [0001] The invention belongs to the field of biological technology, and relates to a genetic engineering strain and a construction method thereof, in particular to a production method and application of a genetic engineering high-yield strain Streptomyces chromogenes and ε-polylysine. Background technique [0002] ε-polylysine is one of the two natural amino acid homopolymers discovered so far (the other is γ-polyglutamic acid). After the discovery of the first strain of ε-polylysine-producing bacteria, ε-polylysine was obtained through soil screening. -The polylysine-producing bacteria all belong to the genera Streptomyces, Streptoverticillum, Kitasatospora, and Cinnamomum The distribution of ε-polylysine producers was mainly restricted to the filamentous bacteria Streptomycesaceae and ergot fungi. ε-polylysine has a broad antibacterial spectrum, and has inhibitory effects on Gram-positive bacteria, Gram-negative bacteria, fungi and some viruses. It has go...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/21C12N15/53C12N15/60C12N15/52C12N15/76C12N15/66C12P13/02C12R1/525
CPCC12N9/001C12N9/88C12N9/93C12N15/76C12N15/66C12P13/02C12Y103/99001C12Y401/01018C12Y401/01017C12Y603/05004
Inventor 谭之磊董天宇贾士儒李小娜崔建东侯颖王贺莉唐昆鹏
Owner TIANJIN UNIV OF SCI & TECH
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