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Application of cucumber CsWRKY50 gene to strengthening downy mildew resistance of cucumber

A cucumber downy mildew and cucumber technology, applied in the field of crop genetic engineering, can solve the problems of pesticide pollution, yield decline, etc., and achieve the effects of safe application methods, improved resistance, and enhanced resistance

Active Publication Date: 2018-09-04
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to provide the application of a cucumber CsWRKY50 gene in enhancing the resistance of cucumber downy mildew, so as to solve the problems that the current cucumber crops are easily attacked by downy mildew during planting, resulting in decreased yield and serious pollution caused by pesticide application

Method used

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  • Application of cucumber CsWRKY50 gene to strengthening downy mildew resistance of cucumber
  • Application of cucumber CsWRKY50 gene to strengthening downy mildew resistance of cucumber
  • Application of cucumber CsWRKY50 gene to strengthening downy mildew resistance of cucumber

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Experimental program
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Effect test

Embodiment 1

[0020] Cloning of Example 1 Cucumber CsWRKY50 Gene and Construction of Overexpression Vector

[0021] (1) Xintai Mici was selected as the test material for the Huangguang variety. This variety has a strong growth potential, with rooting melons on the 4th to 5th nodes, melon-shaped rod-shaped, well-proportioned, green, with dense spines and longitudinal ribs, but not obvious. Cucumber seeds were sown in a solar greenhouse, and common cultivation and management were carried out. When the plants began to produce melons, the young leaves were taken for quick freezing in liquid nitrogen, and the RNA of the young cucumber leaves was extracted by using the Trizol (purchased from Life Technologies company) extraction solution. cDNA was obtained by reverse transcription using a reverse transcription kit from Thermo Fisher, and stored at -20°C for future use.

[0022] (2) Design amplification primers with restriction sites for the cucumber CsWRKY50 gene. The sequences of the primers an...

Embodiment 2

[0040] Example 2 Genetic transformation of cucumber CsWRKY50 gene and acquisition of transgenic plants

[0041] 1. Agrobacterium-mediated genetic transformation of cucumber genes

[0042] Use the Agrobacterium engineering bacteria LBA4404 containing pBI121-35S::CsWRKY50 overexpression vector to infect cucumber cotyledons, and after screening the differentiation medium and rooting medium, perform PCR detection on the regenerated plants with 35S upstream primers and gene downstream primers . The specific steps are as follows:

[0043] (1) Select plump cucumber seeds and soak them in water for about 1 hour, such as figure 2 As shown in middle A, first sterilize with 75% ethanol on a sterile bench for 30 seconds, then sterilize with 4.0% NaClO for 13 minutes, and finally rinse with sterilized ultrapure water for 3 to 4 times. Seeds were sown on MS medium plates with high-temperature sterilized tweezers, and cultured in the dark for 2 days. After germination and rooting, the c...

Embodiment 3

[0061] Embodiment 3 cucumber downy mildew resistance test

[0062] (1) Acquisition of cucumber downy mildew pathogen:

[0063] The diseased leaves containing the mold layer of downy mildew collected from the greenhouse were washed with water and placed in an incubator at 20°C, 90% relative humidity, and kept moist for 24 hours in the dark. When the gray mold layer grew, The diseased leaves are soaked in water, and the sporangia are brushed into a test tube filled with water with a brush to obtain the isolate. Inoculate the isolate on the leaves of the previously planted cucumber susceptible varieties, place the leaves in a large petri dish with water-soaked filter paper at the bottom, absorb the inoculum with a pipette gun, drop it on the back of the leaves, and place it at 22°C during the day and at night In a light incubator at 18°C, leaves covered with sporangia can be obtained from 4d to 6d. Before inoculation, brush the sporangia with a brush in a tissue culture bottle ...

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Abstract

The invention discloses application of a cucumber CsWRKY50 gene to strengthening the downy mildew resistance of the cucumber, the specific application is to overexpress the cucumber CsWRKY50 gene in acucumber plant, and the nucleotide sequence of the cucumber CsWRKY50 gene is shown as in SEQ ID:1. That overexpressing the CsWRKY50 in the cucumber can obviously improve the resistance of the cucumber to the downy mildew is found for the first time, the experiment proves that the cucumber CsWRKY50 gene is transferred into the cucumber plant as a target gene, the expression of the resistance geneis increased by changing the expression quantity of the cucumber CsWRKY50 gene in the cucumber plant, the resistance of the cucumber to the downy mildew is improved, the character is used in the breeding of the cucumber, the cucumber variety capable of resisting the downy mildew is bred by choosing the cucumber CsWRKY50 gene overexpressed plant through the gene detection method, the application method is safe and reliable, and the application has an excellent application prospect in the researches of the resistant variety of the cucumber.

Description

technical field [0001] The invention relates to the technical field of crop genetic engineering, in particular to the application of a cucumber CsWRKY50 gene in enhancing cucumber downy mildew resistance. Background technique [0002] Cucumber (Cucumis sativus L.) is a herbaceous plant of Cucurbitaceae, which is a vegetable crop widely cultivated in the world. Cucumber has many health effects and is deeply loved by people. At present, our country is mainly cultivated in greenhouses and greenhouses. Due to the special climatic conditions and continuous cropping in greenhouses and greenhouses, cucumbers are vulnerable to various diseases, among which downy mildew, powdery mildew, and gray mold are the most serious. The onset and spread of these diseases are fast, causing a series of problems such as a substantial decline in yield, poor quality, and premature plant aging. In order to solve this problem, people usually rely on spraying a large amount of chemical pesticides to p...

Claims

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Application Information

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IPC IPC(8): C12N15/82C12N15/29A01H5/00A01H6/34
CPCC07K14/415C12N15/8282
Inventor 任仲海栾倩倩刘梦雨刘盼静张振
Owner SHANDONG AGRICULTURAL UNIVERSITY
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