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Method for fixed-point knockout of chitin synthase gene

A technology of chitin synthase and fixed-point knockout, which is applied in the fields of genetic engineering, plant gene improvement, botanical equipment and methods, etc., can solve the problems such as the type and family are not clear, and achieve efficient fixed-point knockout and fast fixed-point The effect of knockout

Inactive Publication Date: 2016-09-14
SICHUAN AGRI UNIV
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Problems solved by technology

For the FGSG_06550 gene that QI previously found to be down-regulated in the gene chip treated with salicylic acid, through in-depth analysis of sequence comparison, it was found that this gene does not belong to any identified chitin synthase genes, and the type and family it belongs to not yet clear

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  • Method for fixed-point knockout of chitin synthase gene
  • Method for fixed-point knockout of chitin synthase gene
  • Method for fixed-point knockout of chitin synthase gene

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Embodiment Construction

[0032] In order to make the object, technical solution and advantages of the present invention clearer, the present invention will be further described in detail below in conjunction with examples of implementation. It should be understood that the specific embodiments described here are only used to explain the present invention, not to limit the present invention.

[0033] According to the strategy of reverse genetics, the present invention obtains the definite mutation of the gene through gene knockout technology; explores the effect of the gene on the mycelial growth, sporulation rate, sexual hybridization, germ tube germination, and toxin synthesis of Fusarium graminearum , the integrity of the cell wall were observed; and the recovery mutant and overexpression mutant were used as verification to complete the functional verification of the gene; further through SA identification, the role of the gene in the metabolism of SA mechanism of Fusarium graminearum was observed; t...

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Abstract

The invention discloses a fast and efficient method for fixed-point knockout of a novel chitin synthase gene; the novel chitin synthase gene has a DNA sequence and a cDNA sequence shown in SEQ ID NO:1. The method adopts a freeze-thawing method for transforming a plasmid in agrobacterium AGL-1; 10000 fusarium graminearum conidiospores and agrobacterium with OD600 of 0.75 are co-cultured on a 0.45 [mu] nylon membrane; through construction of an improved homologous recombination plasmid, the construction time of a vector is shortened; the knockout method can greatly shorten the gene knockout time and increase the homologous recombination rate to 93.3%. Through determination of the content of the knockout mutant chitin synthase, the eighth chitin synthase gene is verified to have a function of regulating the synthesis of the chitin synthase. The novel eighth chitin synthase gene is found out in fusarium graminearum for the first time, a new target gene is provided for scab resistance of wheat, and the fast and efficient gene knockout method is provided for research of functional genomics.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and in particular relates to a method for rapidly and efficiently knocking out chitin synthase gene at a fixed point. Background technique [0002] Scab is a worldwide epidemic disease. Scab not only widely affects the yield and quality of crops such as barley, wheat, rye, bare barley (highland barley), rice and corn, but also causes the affected crops to contain a variety of derived mycotoxins and secondary metabolites, resulting in the The seeds of the infringed crops are contaminated, and people and animals will experience acute poisoning such as vomiting, abdominal pain, and dizziness after eating them. And toxins can persist in the food chain for a long time, such as deoxynivalenol and zearalenone, etc., causing serious harm to food safety and human and animal health. Head blight has a wide range of hazards and has caused serious economic losses in major wheat crop producing areas s...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/54C12N15/74C12N15/82
CPCC12N9/1051C12N15/743C12N15/8282C12N2800/101C12N2840/002C12Y204/01016
Inventor 祁鹏飞张亚洲江千涛魏育明郑有良
Owner SICHUAN AGRI UNIV
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