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Method for acquiring diseased plant infected by four pathogens of Yunnan tobacco bushy top disease through injection vaccination

A Yunnan tobacco, injection inoculation technology, applied in the field of plant diseases, can solve the problem of inoculation of pathogens and achieve the effect of enriching preservation methods

Inactive Publication Date: 2018-12-14
YUNNAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to overcome the deficiencies in the background technology, the present invention proposes a method for obtaining infected strains infected with four pathogens of tobacco bushy top disease in Yunnan by injection and inoculation, which can efficiently inoculate four kinds of pathogens of tobacco bushy top disease in Yunnan at the same time. On the plants, it breaks through the technical barrier that the four pathogens of tobacco bushy top disease in Yunnan cannot be inoculated on healthy plants through friction inoculation, avoiding the limitation that the virus must be obtained from living plants when using aphids to transmit viruses and the difference in transmission efficiency due to aphids. Defects that cause experimental errors enrich the preservation of poisonous plant tissues required for inoculation

Method used

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  • Method for acquiring diseased plant infected by four pathogens of Yunnan tobacco bushy top disease through injection vaccination
  • Method for acquiring diseased plant infected by four pathogens of Yunnan tobacco bushy top disease through injection vaccination
  • Method for acquiring diseased plant infected by four pathogens of Yunnan tobacco bushy top disease through injection vaccination

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Effect test

Embodiment 1

[0023] A method for obtaining four kinds of pathogenic strains infected with tobacco bushy top disease in Yunnan by injection and inoculation, the specific steps are:

[0024] Weigh 0.5 g of the plant tissue (fresh sample) containing the four pathogens of tobacco bushy top disease in Yunnan into a sterilized mortar, freeze and grind it into powder with liquid nitrogen, and immediately add 2 mL of PBS buffer solution pre-cooled on ice Grind and mix well, take the grinding solution in a 2mL sterilized centrifuge tube, centrifuge at a medium speed (8000rpm) at 4°C for 3min to obtain the supernatant inoculum, transfer the supernatant inoculum to a new 2mL sterilized centrifuge tube and put it on ice until Subsequent use. After each healthy plant grows to 4 true leaves at the seedling stage, select 2 true leaves at the base for inoculation, use a syringe needle to make 2 holes symmetrically on the main vein at a distance of 2cm, and place your finger on the front of the leaf when i...

Embodiment 2

[0026] A method for obtaining four kinds of pathogenic strains infected with tobacco bushy top disease in Yunnan by injection and inoculation, the specific steps are:

[0027] Weigh 0.5 g of the plant tissues containing the four pathogens of tobacco bushy top disease in Yunnan (frozen samples are stored in a low temperature state of -20°C to -80°C), put them in a sterilized mortar, freeze and grind them into powder with liquid nitrogen Immediately after that, add 2 mL of pre-cooled PBS buffer solution on ice to grind and mix well, take the grinding liquid in a 2 mL sterilized centrifuge tube, centrifuge at a medium speed (8000 rpm) at 4 °C for 3 min to obtain the supernatant inoculum, and transfer the supernatant inoculum to A new 2mL sterilized centrifuge tube was placed on ice for subsequent use. After each healthy plant grows to 4 true leaves at the seedling stage, select 2 true leaves at the base for inoculation, use a syringe needle to make 2 holes symmetrically on the ma...

Embodiment 3

[0029] A method for obtaining four kinds of pathogenic strains infected with tobacco bushy top disease in Yunnan by injection and inoculation, the specific steps are:

[0030]Weigh 0.5 g of the plant tissue containing the four pathogens of tobacco bushy top disease in Yunnan (the dry-preserved sample was dehydrated in a glass desiccator), put it in a sterilized mortar, freeze and grind it into powder with liquid nitrogen, and immediately add 2 mL of it to the ice Grind and mix the pre-cooled PBS buffer solution, put it on ice for 30min, take the grinding solution into a 2mL sterilized centrifuge tube, centrifuge at a medium speed (8000rpm) at 4°C for 3min to obtain the supernatant inoculum, and transfer the supernatant inoculum Transfer to a new 2mL sterilized centrifuge tube and place on ice for subsequent use. After each healthy plant grows to 4 true leaves at the seedling stage, select 2 true leaves at the base for inoculation, use a syringe needle to make 2 holes symmetric...

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Abstract

The invention relates to a method for acquiring a diseased plant infected by four pathogens of a Yunnan tobacco bushy top disease through injection vaccination, belonging to the technical field of plant diseases. The method comprises the following concrete steps: weighing 0.5 g of plant tissues carrying four pathogens of the Yunnan tobacco bushy top disease, grinding the plant tissues into powderwith liquid nitrogen, then adding a PBS buffer solution, carrying out grinding, and centrifuging an obtained homogenate for subsequent injection; injecting the homogenate onto a plant leaf by pricking; in half a month after injection, observing symptoms, and carrying out RT-PCR detection so as to be able to obtain a living sample infected by four pathogens of the Yunnan tobacco bushy top disease.The method provided by the invention can efficiently and simultaneously vaccinate four pathogens of the Yunnan tobacco bushy top disease onto a healthy plant, breaks through the technical barrier thatfour pathogens of the Yunnan tobacco bushy top disease can not be vaccinated onto the healthy plant by friction vaccination, avoids a limitation of needing to acquire viruses from a living plant forvirus transmission with aphids and a defect of causing an experimental error due to an efficiency difference for virus transmission with the aphids, and enriches the preservation manner of virus-carrying plant tissues required for vaccination.

Description

technical field [0001] The invention belongs to the technical field of plant diseases, and in particular relates to a method for obtaining four kinds of pathogenic strains infected with Yunnan tobacco bushy top disease by injection and inoculation. Background technique [0002] Tobacco bushy top disease (TBTD) first occurred in the Chunkewangli tobacco area in northern Zimbabwe in 1957, and then it broke out and caused great losses. In 1993, a large-scale epidemic broke out in Baoshan, Dali and other places in Yunnan. In view of the differences in pathogens, the TBTD found in Yunnan is called Yunnan Tobacco Bushy Top Disease (YTBTD). So far, except for Baoshan, Dali, and Chuxiong, the diseased areas include Kunming, Yuxi, Honghe, Pu'er, Wenshan, Xishuangbanna and Nujiang also had sporadic occurrences of YTBTD. [0003] YTBTD is composed of Tobacco bushy top virus (TBTV), Tobacco bushy top virus satellite RNA (TBTV-SatRNA), Tobacco bushy top virus satellite RNA (TBTV-SatRNA...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01G7/06
CPCA01G7/06
Inventor 李凡马龑刘芳李涛吕婧谭松涛刘琴利李月月肖啟海陈海如
Owner YUNNAN AGRICULTURAL UNIVERSITY
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