Gene cluster affecting the synthesis of Wuyiencin and the method and application of screening and identifying the gene cluster

Abstract

The invention provides a gene cluster 26 affecting synthesis of wuyiencin. A method for rapidly screening and identifying the gene cluster with microbial functions comprises the following steps that firstly, target genes are selected; secondly, a recombined gene carrier lack of the target genes are built through a Gibson method, specifically, (1) a primer of the target genes is designed for amplifying two homologous arms, and an overlap region is arranged on the primer; (2) PCR amplification is conducted on the sequence of the primer in the step (1), and the left homologous arm and the right homologous arm are obtained; (3) restriction enzyme cutting sites of a plasmid are selected to cut the plasmid, and thus a linear plasmid is obtained; (4) the left homologous arm and the right homologous arm in the step (2) are assembled on the linear plasmid, and an assembled product is obtained; and (5) the assembled product is used for building the recombined gene carrier in large quantity; and thirdly, the recombined gene carrier is used for building a mutant strain, and whether the target genes are functional genes or not is verified. The method for screening and identifying the gene cluster and application has the beneficial effects that the time of building the recombined gene carriers through the method is shortened, and the select breeding progress of the strain genes is accelerated.

Description

technical field [0001] The invention relates to the field of biotechnology, and more specifically relates to a gene cluster affecting the synthesis of Wuyiencin and a method and application for screening and identifying the gene cluster. Background technique [0002] Biologically active natural products (secondary metabolites) produced by microorganisms, which are human resistance to cancer, aging and infectious diseases, anticancer drugs, clinical and agricultural antibiotics, and immunosuppressants, etc., can be widely used in humans, animals and Agriculture and other aspects have important application value for human production and life. The genomes of microorganisms that have been sequenced also contain a large number of invisible natural product biogenesis gene clusters. This information provides a valuable resource library for the screening and mining of new functional natural drugs in the future. The traditional method of library construction and screening is cumberso...

AI Technical Summary

Problems solved by technology

However, in the process of constructing recombinant vectors, there are many disadvantages in the traditional enzyme-cut ligation, for example: repeated enzyme-cut ligation and verification are required; when assembling multiple fragments, multiple different enzyme-cut sites need to be involved; The T vector is cumbersome as an intermediate vector; the connection efficiency is low when the assembly fragment is long

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