Cloning and sequence analyzing method for active product gene capable of resisting cucurbits fusarium wilt

Abstract

The invention belongs to the technical field of cucurbits fusarium wilt prevention and cure, and discloses a cloning and sequence analyzing method for an active product gene capable of resisting cucurbits fusarium wilt. The cloning and sequence analyzing method for the active product gene capable of resisting the cucurbits fusarium wilt comprises the steps that soil microbial genome DNA is extracted through an ectopic cracking method; metagenome Fosmid library construction and quality detection thereof are conducted; a metagenome Fosmid expression library is screened, and Fosmid active cloning is obtained; active substances are purified and authenticated, the structures of the active substances are determined, and the activity of a compound resisting the cucurbits fusarium wilt is verified; and plant endogenous strains are screened and transformed, genes are converted into the endogenous strains which are subjected to transforming, and a high-efficiency expression system of drug-related genes resisting the cucurbits fusarium wilt is established by selecting different promoters, different culture media and fermentation conditions. According to the cloning and sequence analyzing method for the active product gene capable of resisting the cucurbits fusarium wilt, the plant endogenous strains which are separated, screened and transformed serve as an expression strain system, and a target gene heterogenous high-efficiency expression system which is screened from a library and has the biological activity is established. A solid foundation is laid for commercialization of drugs resisting the fusarium wilt.

Description

Technical field [0001] The invention belongs to the technical field of prevention and control of cucurbit wilt, and particularly relates to a method for cloning and sequence analysis of a gene of the active product resistant to cucurbit wilt. Background technique [0002] Melon wilt, also known as vine cut disease and wilt disease, is one of the main soil-borne diseases in the production of melons, especially watermelons, cucumbers and melons. The disease spreads to dozens of countries and regions including China, the United States, Italy, Israel, Japan, and India, and watermelon growing areas in the north and south of my country have occurred to varying degrees. Especially in the southern tropical and subtropical areas, where there is abundant rainfall and mild climate, pathogenic bacteria have no overwintering phenomenon, resulting in a large accumulation of pathogenic spores in the soil, and the incidence of disease is extremely high. The general incidence of repeated cropping...

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Problems solved by technology

[0003] In summary, the existing problems in the prior art are: at present, most of them are controlled by chemical agents. Since Fusarium wilt is a soil-borne disease, its pathogenic bacteria have a strong masking ability, and it is difficult for chemical fungicides to directly contact with it, so it cannot be fundamentally killed. The cost of application is high to kill pathogenic bacteria; at the same time, a series of side effects such as polluting the environment and destroying ecological balance caused by the widespread use of chemical agents have become increasingly prominent, and production and use are gradually being restricted; in addition, long-term use of chemical drugs, pathogenic bacteria are prone to drug resistance , over time, chemical drugs lose their original effect

The probability of success in discovering new antibiotic drugs is getting smaller and smaller, and there is often a phenomenon of repeated screening

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