Seedling-stage injection inoculation method for zucchini cucumber mosaic virus

A cucumber mosaic virus, injection inoculation technology, applied in biochemical equipment and methods, microbe measurement/inspection, etc., can solve the problems of late inoculation time, unfavorable identification, and inability to clarify pathogens, etc., to achieve fast growth and shorten Time, good effect of resistance

Pending Publication Date: 2021-03-23
华盛农业集团股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because zucchini virus disease is caused by a variety of pathogens, the impact on the inoculation results is relatively complicated, and the role of the main pathogen cannot be clarified, and it is not conducive to repeated identification, which affects the consistency and objectivity of the inoculation results. directly affect the reliability of vaccination results
In the prior art, there has been a seedling-stage inoculation method for pepper cucumber mosaic virus disease, which injects the cotyledons with the virus at the stage of two true leaves of pepper seedlings, but this method is not suitable for zucchini plants. There are large differences in growth rate and shape, and the time of inoculation in the above-mentioned prior art is relatively late, so it is difficult to meet the requirements of disease resistance identification

Method used

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  • Seedling-stage injection inoculation method for zucchini cucumber mosaic virus
  • Seedling-stage injection inoculation method for zucchini cucumber mosaic virus
  • Seedling-stage injection inoculation method for zucchini cucumber mosaic virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1 The incidence of CMV in zucchini seedlings by friction inoculation and injection inoculation

[0028] The inoculation method used was as follows:

[0029] (1) After the zucchini seeds were sterilized in 2% sodium hypochlorite for 10 minutes, rinse them with sterile water for 3-4 times;

[0030] (2) Zucchini seeds were sterilized and germinated in a Petri dish at 30°C in the dark for 36 hours. After the seeds germinate, they are sown in a 7×7 nutrient pot for growth. During the growth period, the temperature is 25°C day / night 20°C, and the seedlings can be inoculated when 2 cotyledons are fully unfolded. Specific forms such as figure 1 ;

[0031] (3) Take the purified cucumber mosaic virus disease bacterial strain, artificially inject and inoculate it on cotyledon flattening stage zucchini seedlings, and harvest the diseased leaves three weeks after the inoculation; have both;

[0032] (4) Add 2.5 ml of 0.01 mol / L phosphate buffer solution with a pH value ...

Embodiment 2

[0037] Example 2 RT-PCR detection of field-onset zucchini virus disease infestation species

[0038] 1. Take tender leaves from the zucchini leaves of each plant obtained in Example 1, and store them at -80°C after rapid liquid nitrogen freezing;

[0039] 2. Extraction of total RNA

[0040]Weigh 0.5g of each sample, put them into a -80°C deep-frozen mortar and add liquid nitrogen to fully grind, quickly transfer to a sterile 1.5mL Eppendorf tube pre-cooled by liquid nitrogen, and add 1mL Trizol pre-cooled at 4°C for extraction Mix well and let stand for 5 minutes, then centrifuge at 12,000g for 5 minutes at 4°C. The supernatant was extracted once with 200 μL chloroform, and centrifuged at 12,000 g for 15 min at 4°C. Add an equal volume of isopropanol to the supernatant, invert and mix well, let stand for 10 min, and centrifuge at 12,000 g for 15 min at 4°C. Discard the supernatant, wash the pellet once with 70% pre-cooled ethanol, and dissolve it in 30 μL DEPC-treated ddH ...

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Abstract

The invention relates to the field of plant virus inoculation, in particular to a seedling-stage injection inoculation method for zucchini cucumber mosaic virus. The method adopts a needle tube injection mode, and can be used for inoculation when zucchini plants are cultured until cotyledons are completely unfolded, compared with the existing needle tube injection method, the inoculation time is greatly advanced, and the condition of the zucchini seedlings can be investigated 10 days after inoculation. According to the method, the virus inoculation amount is easy to control, damage to leaves is small and is closer to the field morbidity situation, inoculation liquid can be directly injected into mesophyll cells, observation is easy, and missed inoculation is reduced.

Description

technical field [0001] The invention relates to the field of plant virus inoculation, in particular to a seedling stage injection inoculation method for zucchini cucumber mosaic virus disease. Background technique [0002] Zucchini is one of the bulk vegetables in the world, and it is also the main melon vegetable in my country. The annual zucchini planting area is about 5 million mu or more, among which Shandong, Shanxi, Yunnan, Northwest and other regions are important high-quality zucchini producing areas in my country. Zucchini virus disease is a common and harmful disease. There are three main viruses infecting zucchini: Cucumber mosaic virus (CMV), Watermelon mosaic virus (WMV) and Zucchini yellow mosaic virus (ZYMV). Cucumbermosaicvirus (CMV) is one of the most important pathogens causing zucchini virus disease. After CMV infects zucchini, it shows a gradual pathogenesis. In the early stage of the disease, the base of young leaves turns yellow, and then the yellow...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/18
CPCC12Q1/18
Inventor 袁晓伟李兴盛蒋新萍李英杰
Owner 华盛农业集团股份有限公司
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