Method for improving extracellular pigment of Monascus
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A technology of Monascus mold and pigment, applied in the field of genetic engineering, can solve the problem of low yield of extracellular pigments of Monascus
Active Publication Date: 2019-01-18
CENTRAL SOUTH UNIVERSITY OF FORESTRY AND TECHNOLOGY
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[0004] The technical problem to be solved in the present invention is to provide a method for improving the extracellular pigment of Monascus to
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Embodiment 1
[0041] 1. Construction of the binary plasmid expression vector of Monascus_07017 gene deletion mutant strain of Monascus purple
[0042] (1) using software to analyze the sequence shown in SEQ ID No: 1;
[0043] (2) Using the cDNA of Monascus purple as a template, 7017-up-osc-F (sequence is SEQ ID No: 2), 7017-up-osc-R (sequence is SEQ ID No: 3) as a pair Primer and 7017-dn-osc-F (sequence is SEQ ID No: 4), 7017-dn-osc-R (sequence is SEQ ID No: 5) are a pair of primers to carry out PCR amplification respectively to obtain the 5' of Monascus_07017 gene and 3' homology arm fragments, see figure 2 ;
[0044] (3) Plasmid pXS (donated by the research group of Professor Liu Tiangang of Wuhan University, see the plasmid map for details figure 1 ) as a template, G418-osc-F (sequence is SEQ ID No: 6), G418-osc-R (sequence is SEQ ID No: 7) as a pair of primers for PCR amplification to obtain the G418 gene fragment.
[0045] (4) Using Overlap PCR technology, using the 5' and 3' homo...
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Abstract
The invention provides a method for improving extracellular pigment of Monascus. The method comprises knocking out gene encoding ERG4/ERG24 family protein on ergosterol synthesis pathway in Monascus.ERG4/ERG24 gene encode in Monascus are knocked out by an agrobacterium-mediated transformation technology. As that plant binary plasmid pCAMBIA1300 carry is used, the recombinant plasmid is mediate byAgrobacterium tumefaciens EHA105 to be transformed into Monascus, the production level of extracellular pigment is remarkably improved, and the result shows that the recombinant strain has good passage stability. A liquid fermentation result show that that extracellular monascus pigment of the parent strain is 4.92 U/mL, the extracellular monascus pigment of the recombinant strain constructed bythe invention is 6.52 U/mL, and the production amount of the extracellular pigment is increase by 32.52%, which shows that the production amount of the extracellular pigment of the monascus can be remarkably increased by the method of the invention.
Description
technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a method for increasing the extracellular pigment of monascus. Background technique [0002] Monascus, as a filamentous fungus, belongs to the kingdom Fungi, phylum Ascomycota, class Ascomycetes, order Phytomycota, and family Monascus. Monascus pigment is a series of natural pigments of polyketides produced by the secondary metabolism of Monascus fungus. It has been widely used as food additives in beverages, sauces, edible oils, bread, pastries and other fields in China, Korea and Japan. It has a history of thousands of years. At the same time, because the red yeast pigment 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-bacteria, anti-fungal, etc., it The applic...
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