Gene cluster expression strengthened recombinant streptomyces diastatochromogenes and construction method thereof

Abstract

The invention discloses a gene cluster expression strengthened recombinant streptomyces diastatochromogenes and a construction method thereof. The gene cluster expression strengthened recombinant streptomyces diastatochromogenes excessively expresses a whole toy gene cluster for biosynthesizing toyocamycin and has stronger toyocamycin synthetic capability than streptomyces diastatochromogenes 1628. A construction process comprises the following steps: 1) constructing an expression vector pSET152-toyAM; and 2) integrating the expression vector into chromosome of the streptomyces diastatochromogenes by utilizing a conjugative transferring method to obtain an engineering bacterium. According to the construction method disclosed by the invention, specificity of the vector pSET152-toyAM is integrated into the chromosome of the streptomyces diastatochromogenes 1628 by utilizing the conjugative transferring method to obtain an engineering bacterium with genetic stability. Compared with an original strain, the gene cluster expression strengthened recombinant streptomyces diastatochromogenes has the advantage that the yield of toyocamycin is increased by 4.5 times.

Description

technical field [0001] The invention relates to improving the yield of toyocamycin by strengthening the expression of gene cluster toy AM in amylase chromogenic streptomyces, and belongs to the technical field of genetic engineering. Background technique [0002] Toyocamycin is a new type of nucleoside antibiotics with the molecular formula C 12 h 13 N 5 o 4 , Ribose C 1 The deazapurine ring similar to guanine is connected, and the core structure is a pyrropyrimidine nucleoside analogue. The mechanism of action is mainly to affect the growth of bacteria by inhibiting the transcription of microorganisms, and the research reports on its biological activity are mainly concentrated in the field of clinical medicine. Recent studies have found that toyocamycin has a good control effect on a variety of plant diseases, long-term use will not cause environmental pollution, and it also has a certain regulatory effect on plant growth. Therefore, toyocamycin has great application ...

AI Technical Summary

Problems solved by technology

In the previous study, our research group used the toyocamycin-producing strain Streptomyces chromogenes (Streptomyces chromogenes) with independent intellectual property rights Streptomyces diastatochromogenes ) 1628 as the research object, cloned some structural genes in the gene cluster, such as toyB, toyF, etc., and tried to S. diastatochromogenes overexpressed in 1628 and also cloned S. diastatochromogenes The pleiotropic regulatory factor adpA responsible for secondary metabolism in 1628, but the increase in the yield of toyocamycin is only 20%-30%, which still needs to be improved for industrial production

Using metabolic engineering technology to increase the copy number of the original strain biosynthetic toyocamycin gene cluster instead of a single structural gene, thereby increasing the metabolic flux of toyocamycin synthesis and increasing the production of toyocamycin, because the problems involved are complex and there is no correlation to report

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