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Genetic transformation method of maize curvularia germination conidia

A technology of conidia and genetic transformation, which is applied in the field of genetic transformation of germination conidia of C. good effect

Inactive Publication Date: 2009-03-11
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since the germ tubes of the germinated spores are attached to the membrane pores at this time, it is difficult to wash the germinated spores from the membrane. At the same time, the washed spore liquid is mixed with many other impurities, which are not easy to remove. Low efficiency and poor repeatability

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] 1. Preparation of germinated conidia of Curvularia maize

[0017] On the potato-glucose-agar medium, after Curvularia maize was cultivated for 7 days, the conidia of Curvularia maize was scraped off with 0.9% normal saline and then filtered with sterile lens paper to remove the hyphae, and the obtained cornbent Spore conidia filtrate was adjusted to a concentration of 10 with 0.9% saline 8 Spores / ml; Take a Petri dish with a layer of water-retaining filter paper, add sterile water to wet the filter paper and form a water film on the surface, add two wooden sticks on it, and then take a concentration of 10 8 Put 200 μL of spores / ml Curvularia maize conidia suspension in the center of a sterile and clean glass slide, divide it into 4 drops, then quickly turn the slide upside down, and place the two ends of it in a petri dish On two wooden sticks, cultivate in the dark at 30°C for 9 hours; the germinated conidia of Curvularia maize were washed with 0.9% normal saline, cen...

Embodiment 2

[0023] 1. Preparation of germinated conidia of Curvularia maize

[0024]On the potato-glucose-agar medium, after Curvularia maize was cultivated for 7 days, the conidia of Curvularia maize was scraped off with 0.9% normal saline and then filtered with sterile lens paper to remove the hyphae, and the obtained cornbent Spore conidia filtrate was adjusted to a concentration of 10 with 0.9% saline 6 Spores / ml; Take a Petri dish with a layer of water-retaining filter paper, add sterile water to wet the filter paper and form a water film on the surface, add two wooden sticks on it, and then take a concentration of 10 6 Put 200 μL of spores / ml Curvularia zea conidia suspension in the center of a sterile and clean glass slide, divide it into 3 drops, then quickly turn the slide upside down, and place the two ends of it in a petri dish On two wooden sticks, cultivate in the dark at 28°C for 6 hours; the germinated conidia of Curvularia maize were washed with 0.9% normal saline, centri...

Embodiment 3

[0030] 1. Preparation of germinated conidia of Curvularia maize

[0031] On the potato-glucose-agar medium, after Curvularia maize was cultivated for 7 days, the conidia of Curvularia maize was scraped off with 0.9% normal saline and then filtered with sterile lens paper to remove the hyphae, and the obtained cornbent Spore conidia filtrate was adjusted to a concentration of 10 with 0.9% saline 7 Spores / ml; Take a Petri dish with a layer of water-retaining filter paper, add sterile water to wet the filter paper and form a water film on the surface, add two wooden sticks on it, and then take a concentration of 10 7 Put 200 μL of spores / ml Curvularia maize conidia suspension in the center of a sterile and clean glass slide, divide it into 4 drops, then quickly turn the slide upside down, and place the two ends of it in a petri dish On two wooden sticks, cultivate in the dark at 30°C for 3 hours; the germinated conidia of Curvularia maize were washed with 0.9% normal saline, cen...

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Abstract

The invention relates to a method for genetic transformation of germinating conidiophore of Curvularia lunata Boed. The method comprises the following steps: culturing the germinating conidiophore of the Curvularia lunata Boed as a receptor by a slide suspension method; co-culturing agrobacterium containing an epiphyte conversion vector system and the germinating conidiophore; and screening transformants with resistibility on an antibiotic culture medium. The system uses the Curvularia lunata Boed as initial bacteria, spore is scratched by 0.9 percent of physiological saline after the Curvularia lunata Boed is cultured on the PDA culture medium for 5 to 7 days, and is used for infection of the agrobacterium after the spore is germinated by the slide suspension culture method. The infected germinating conidiophore obtains resistant transformant by co-culture, bacteriostasis culture and anti-selective agent screening culture of continuous two generations, so that a Curvularia lunata Boed mutant base is constructed to provide rich screening strain sources for weak pathogenicity and pathogenic defective Curvularia strain, and meanwhile can be used for cloning related gene of Curvularia lunata Boed pathogenicity so as to lay a foundation for further investigating the pathogenesis mechanism of the Curvularia lunata Boed strain.

Description

technical field [0001] The invention relates to a method for genetic transformation of germinated conidia of Curvularia maize, in particular to a method for genetic transformation of germinated conidia mediated by Agrobacterium and using the germinated conidia cultured by the slide suspension method as a material to obtain cornbent A method for a spore mutant belongs to the field of biotechnology. Background technique [0002] Curvularia lunata (Wakker) Boed, caused by Curvularia lunata (Wakker) Boed, has caused major damage in most countries of the world. In 1996, the disease caused severe maize loss in China, and 40% of the maize production areas were infected by the disease. In order to better control the disease and breed resistant varieties, it is of great significance to conduct in-depth research on the pathogenic molecular mechanism of the pathogen. Isolating and identifying pathogenicity-related genes of Curvularia sp. is the key to achieve this goal, and creating ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14C12N3/00C12R1/645
Inventor 陈捷刘铜刘力行姜雪
Owner SHANGHAI JIAO TONG UNIV
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