A method for knocking out negative regulatory protein genes to increase the level of acarbose fermentation

A technology of acarbose and fermentation level, which is applied in the field of biomedicine to achieve the effect of improving transcription level and improving fermentation yield

Active Publication Date: 2022-08-09
SHANGHAI JIAO TONG UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, there are still few studies involving the relevant transcriptional regulators and their regulatory mechanisms in the acarbose biosynthetic pathway

Method used

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  • A method for knocking out negative regulatory protein genes to increase the level of acarbose fermentation
  • A method for knocking out negative regulatory protein genes to increase the level of acarbose fermentation
  • A method for knocking out negative regulatory protein genes to increase the level of acarbose fermentation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0039] This example is a specific process for obtaining mutant strains WXM-06 and WXM-07 that knock out the negative regulatory protein genes ACPL_5445 and ACPL_3989. The specific operation steps are as follows:

[0040] Step 1: Construction of plasmid pLQ1455

[0041] Taking the genomic DNA of Actinomyces mobilis SE50 / 110 as the template, using the primer ACPL_5445-LF / LR, the 1474bp gene fragment on the left side of ACPL_5445 was obtained by PCR amplification, and the 1646bp gene fragment on the right side of ACPL_5445 was obtained by PCR amplification using the primer ACPL_5445-RF / RR Gene fragment, the correctness of the target gene is confirmed by gene sequencing. The 1474 bp PCR fragment from the left side of the ACPL_5445 gene and the 1646 bp PCR fragment from the right side of the ACPL_5445 gene were inserted into the XbaI / HindIII site of the plasmid pLQ752 to obtain the plasmid PLQ1455. Under the condition of 37°C, two kinds of restriction enzymes, XbaI and HindIII, w...

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Abstract

The invention discloses a method for knocking out a negative regulation gene to improve the fermentation level of acarbose. By adopting the method of the invention, by knocking out the negative regulatory protein genes ACPL_5445 and ACPL_3989 respectively in the actinomycete mobilis, a mutant strain with high acarbose production is obtained. By knocking out the negative regulatory protein gene in the present invention, the expression of the acarbose biosynthesis gene can be promoted, and finally the acarbose production can be significantly improved. Compared with the starting strain, the fermentation yield of the high-yielding strain obtained by the present invention is increased by 13% and 8% respectively, and the fermentation level of the laboratory shake flask reaches 3.62g / L and 3.47g / L respectively.

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to a method for knocking out negative regulation genes to improve the fermentation level of acarbose; Bose biosynthesis gene transcription levels, thereby increasing acarbose production. Background technique [0002] Currently, the number of people with diabetes continues to increase worldwide. Type 2 diabetes is the most common type of diabetes, accounting for more than 90% of all diabetes. 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 structure control and moderate exercise are effective intervention methods. The classic hypoglycemic drugs currently used for the treatment of diabetes mainly include insulin secretagogues, insulin sensitizers, and α-glucosidase inhibitors. [0003] Acar...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/21C12N15/31C12N15/74C12P19/26C12R1/045
CPCC07K14/195C12N1/20C12N15/74C12P19/26
Inventor 白林泉汪雪梅
Owner SHANGHAI JIAO TONG UNIV
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