Construction method of high yield monascorubin strain

A technology of high-yield strains and construction methods, applied in the direction of using vectors to introduce foreign genetic material, recombinant DNA technology, etc., can solve the problem of low yield of monascus pigment

Active Publication Date: 2019-02-22
JIANGXI SCI & TECH NORMAL UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] The present invention aims at the technical defects of the prior art, and provides a method for constructing a high-yielding strain o

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  • Construction method of high yield monascorubin strain
  • Construction method of high yield monascorubin strain
  • Construction method of high yield monascorubin strain

Examples

Experimental program
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Effect test

Embodiment 1

[0041] 1. Construction of binary plasmid knockout vector pHph0380.

[0042] The original vector was the commercial plant binary plasmid pCambia0380. Design a pair of oligonucleotide sequences F&R, which were synthesized by Shanghai Sangon Bioengineering Co., Ltd. This sequence in turn contains the following restriction endonuclease sites (Hind III, Kpn I, Sac I, Pac I, Pme I, Xho I, Xba I, Bgl II). The binary plasmid vector pCambia0380 was cut with restriction endonucleases Hind III and Bgl II. The oligonucleotide sequence was ligated with the restriction vector pCambia0380 by T4 DNA ligase to obtain the binary plasmid vector pCambia0380G.

[0043] Using the plasmid pMD19-PgpdA-hph-TtrpC preserved in the laboratory as a template, the hph expression cassette was amplified with primers PgpdA-Sac I-F&TtrpC-Xho I-R. The hph expression cassette fragment and the binary plasmid vector pCambia0380G were simultaneously digested with restriction endonucleases Sac I and Xho I. The hp...

Embodiment 2

[0096] 1. Construction of binary plasmid knockout vector pHph0380

[0097] The original vector was the commercial plant binary plasmid pCambia0380. Design 1 pair of oligonucleotide sequences F&R, containing in order the following restriction endonuclease sites (Hind III, Kpn I, Sac I, Pac I, Pme I, Xho I, XbaI, Bgl II). The binary plasmid vector pCambia0380 was cut with restriction endonucleases Hind III and Bgl II. The oligonucleotide sequence was connected to the vector by T4 DNA ligase to obtain the binary plasmid vector pCambia0380G.

[0098] Using the plasmid pMD19-PgpdA-hph-TtrpC preserved in the laboratory as a template, the hph expression cassette fragment was amplified with primers PgpdA-Sac I-F&TtrpC-Xho I-R. The promoter fragment and the binary plasmid vector pCambia0380G were digested simultaneously with restriction endonucleases Sac I and Xho I. The oligonucleotide sequence was connected to the vector by T4 DNA ligase to obtain the binary plasmid knockout vecto...

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Abstract

The invention provides a construction method of a high yield monascorubin strain. The method comprises the steps of connecting a pair of oligonucleotide sequences sequentially comprising Hind III, KpnI, Sac I, Pac I, Pme I, Xho I, Xba I and Bgl II enzyme cutting sites to dual plant plasmid pCambia0380 to construct a double plasmid expression vector pCambia0380G, connecting an hph expression cassette segment with the expression vector to form a double plasmid knockout vector pHph0380, amplifying upstream and downstream homologous arm segments of a gltp1 gene, connecting the homologous arm segments to the double plasmid knockout vector pHph0380 to form a double plasmid knockout vector pHph0380-GLTP, mediating and transforming the vector pHph0380-GLTP from a root knot agrobacterium EHA105 toparent monascus ruber CICC41233 to accomplish construction of the high yield strain. Experimental verification shows that the overall productivity of monascorubin is improved, an alcohol soluble pigment can be accumulated directionally, and at the same time, the accumulation of the monascorubin in a fermentation process is advanced significantly.

Description

technical field [0001] The invention relates to the technical field of industrial microbes, and further relates to genetic engineering technology and mold fermentation technology, in particular to a method for constructing a high-yield strain of monascus pigment. Background technique [0002] Monascus pigment is a natural pigment fermented by microorganism Monasus spp. with rice as raw material. It has a history of more than one thousand years in my country. As a food additive, monascus pigment is widely used in food processing and cosmetics manufacturing and other fields; because it also has a wide range of biological activities such as regulating blood lipids, lowering blood pressure, preventing vascular sclerosis, anti-diabetes, inhibiting obesity, anti-inflammation, anti-allergic, anti- Peroxidation, anti-cancer, anti-bacterial, anti-fungal, etc., its application in the development of probiotics and health care products and the medical field has also attracted more and m...

Claims

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Application Information

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IPC IPC(8): C12N15/74C12N15/66
CPCC12N15/66C12N15/74
Inventor 龙传南曾斌陶琴琴刘心怡刘梦梦彭玲程芳婷王淑琴吾蔚蔚
Owner JIANGXI SCI & TECH NORMAL UNIV
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