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Method for enhancing positive regulatory protein gene expression to increase acarbose fermentation level

A kind of acarbose, fermentation level technology, applied in the field of bioengineering, can solve problems such as no research findings

Active Publication Date: 2021-04-02
SHANGHAI JIAO TONG UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

So far, there are still few studies on the relevant transcriptional regulators and their regulatory mechanisms in the acarbose biosynthetic pathway, and no research has found that there are genes that regulate the transcription of the synthetic pathway in the acb gene cluster

Method used

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  • Method for enhancing positive regulatory protein gene expression to increase acarbose fermentation level
  • Method for enhancing positive regulatory protein gene expression to increase acarbose fermentation level
  • Method for enhancing positive regulatory protein gene expression to increase acarbose fermentation level

Examples

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Embodiment

[0047] This example is the specific process of obtaining the enhanced expression mutant strains WXM-01, WXM-02, WXM-03, WXM-04, WXM-05 of the positive regulatory genes ACPL_1889, ACPL_4236, ACPL_7303, ACPL_6479 and ACPL_8104. The specific operation steps are as follows:

[0048] Step 1: Construction of plasmid pLQ1450

[0049] Using the genomic DNA of actinomycetes SE50 / 110 as a template, using primers GBACPL_1889-F / R, the ACPL_1889 gene was amplified by PCR, and the correctness of the target gene was confirmed by gene sequencing; using the pDR3 plasmid as a template, kasOp- F / R, the kasOp* gene was obtained by PCR amplification, and the correctness of the target gene was confirmed by gene sequencing. The amplified fragment after digestion was inserted into the XbaI / BamHI site of plasmid pSET152 to obtain plasmid PLQ1450. Under the condition of 37°C water bath, the target band of about 350bp can be observed by using XbaI and BamHI two restriction endonucleases for enzyme dig...

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Abstract

The invention discloses a method for enhancing positive regulatory protein gene expression to improve the acarbose fermentation level. In actinoplanes, positive regulatory protein genes ACPL_1889, ACPL_4236, ACPL_7303, ACPL_6479 and ACPL_8104 are respectively subjected to intensified expression by utilizing a strong promoter kasOp*, so that an acarbose high-yield mutant strain is obtained. Enhancement on the expression of the positive regulation protein genes can promote the expression of acarbose biosynthetic genes, and finally, obviously improve the acarbose yield. Compared with an originalstrain, the fermentation yields of the high-yield strains obtained by the invention are respectively improved by 25%, 18%, 26%, 22% and 15%, and the laboratory shake flask fermentation levels respectively reach 3.29g / L, 3.11g / L, 3.32g / L, 3.21g / L and 3.03g / L.

Description

technical field [0001] The present invention relates to the field of bioengineering, in particular to a method for enhancing the expression of a positive regulatory protein gene to improve the fermentation level of acarbose; by respectively strengthening the expression of the positive regulatory protein gene in actinomycetes QQ-2 ACPL_1889, ACPL_4236, ACPL_7303, ACPL_6479, and ACPL_8104 can increase the transcription level of acarbose biosynthetic genes, thereby increasing the production of acarbose. Background technique [0002] Diabetes is a metabolic disease characterized by hyperglycemia. Currently, type 2 diabetes is the most common type of diabetes, accounting for more than 90% of the total number of diabetic patients. Many clinical studies have proved that reasonable control of postprandial blood sugar can effectively slow down or reduce the occurrence of some chronic cardiovascular and cerebrovascular complications. Oral hypoglycemic drugs combined with dietary stru...

Claims

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Application Information

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IPC IPC(8): C12N1/21C12N15/74C12N15/31C12P19/26C12R1/045
CPCC07K14/365C12N15/74C12P19/26
Inventor 白林泉汪雪梅
Owner SHANGHAI JIAO TONG UNIV
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