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Heredity transformation method of coix seeds mediated by agrobacterium tumefaciens

An Agrobacterium-mediated and genetic transformation technology is applied in the field of genetic transformation of Coix stem tips mediated by Agrobacterium, which can solve problems such as inability to improve breeding.

Inactive Publication Date: 2020-01-07
GUIZHOU UNIV
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  • Application Information

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Problems solved by technology

At present, the stable genetic transformation system has been widely used in monocotyledonous plants such as corn and wheat, but the application in Coix has not been clearly reported, so we cannot improve and breed it from the means of modern molecular biology

Method used

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  • Heredity transformation method of coix seeds mediated by agrobacterium tumefaciens
  • Heredity transformation method of coix seeds mediated by agrobacterium tumefaciens
  • Heredity transformation method of coix seeds mediated by agrobacterium tumefaciens

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Embodiment 1

[0034] Example 1 A technique of genetic transformation of Coix mediated by Agrobacterium

[0035] 1.1 Seed disinfection: Take an appropriate amount of seeds and use detergent to clean the impurities on the surface, then put them in boiling water for 3-4 seconds, take them out, dry them naturally and soak them in carbendazim containing 1‰ for 7-8 hours. Take it out and wash it, place it on a culture plate, and place it in a constant temperature incubator at 28°C until the seeds germinate, and the stem tip grows to 2-3cm. Use a scalpel to cut the stem tip of the germinated Coix coix seedlings to expose the shoot tip meristem Organized for backup. See figure 1 shown.

[0036] 1.2 Bacterial cultivation: Agrobacterium tumefaciens strain LBA4404, carrier PcambiaU1301 carrier (commercially available) carrying hygromycin resistance gene Hyg and GUS gene was inoculated into a mixture containing 50 mg / L kanamycin (Kan) and 50 mg / L rifampicin (Rif) on YEP solid medium, cultivated in 2...

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Abstract

The invention relates to a heredity transformation method of coix seeds mediated by agrobacterium tumefaciens. Sprouting treatment is performed on mature coix seeds, and transection is performed untilstem tip meristem is a heredity transformation receptor. The method comprises the following specific operation steps of (1) preparing the heredity transformation receptor; (2) culturing agrobacteriumtumefaciens; (3) performing heredity transformation on coix seeds; and (4) transplanting and managing genetically modified plants. The method does not adopt a tissue culture technique, agrobacteriumtumefaciens are cultured, and a sterile operation is not need. The method which is used has the advantages of being simple to operate and low in cost, capable of transforming klenow fragment DNA, being stable in heredity, and low in copy number and the like, and the method becomes a heredity transformation method which is broad to apply.

Description

technical field [0001] The technical field of the invention is plant biotechnology, and specifically relates to a method for using Agrobacterium to mediate the genetic transformation of coix stem tip. Background technique [0002] Job's tears is a medicinal and edible economic crop belonging to the family Gramineae, which has important medicinal value and extremely high nutritional value. In recent years, the barley rice industry in Guizhou Province has developed rapidly, and the province's planting area has reached more than 40,000 hm 2 , accounting for more than 80% of the country's planted area. However, there are problems such as high stem, easy lodging, and poor disease resistance in the small white shell of coix xingren, which affect the increase of its yield and restrict its industrial development. [0003] The Agrobacterium-mediated method has the advantages of simple operation, low cost, ability to transform large fragments of DNA, stable genetics, and low copy nu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/84
CPCC12N15/8205
Inventor 赵懿琛谢梅曾晓芳王兰兰王银赵德刚
Owner GUIZHOU UNIV
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