Method for treating paddy rice tissues by using prokaryotic expression product and inducing generation of callose

A technology of prokaryotic expression and callose, which is applied in chemical instruments and methods, plant cells, peptides, etc., to achieve the effect of simple method and avoiding autofluorescence interference

Inactive Publication Date: 2012-07-11
YUNNAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no report on a method to directly use the prokaryotic expression product of the gene to detect the callose production of the host root tissue induced by the effector protein of rice blast fungus in vitro, so as to quickly identify whether the effector protein gene triggers the basic defense response of the host

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0013] The rice root tissue was treated with 1.0 mg / ml fusion protein for 12 hours, and the formation of callose was directly observed after aniline blue staining.

[0014] Centrifuge the expression product of the effector protein gene of Magnaporthe grisea to collect the cells, and suspend the cells with freshly prepared buffer (20mM Tris-HCl, pH7.4; 200mM NaCl; 1mM EDTA; 10mM β-mercaptoethanol) . The supernatant was collected by centrifugation after ultrasonography to disrupt the cells in an ice-water bath. The treatment is to treat the root tissue of the disease-resistant rice variety Nipponbare at a concentration of 1.0 mg / ml after purification of the prokaryotic expression product of the gene for 12 hours, and directly stain with aniline blue to observe callose. As a control, after the prokaryotic expression product of the gene was purified, the root tissue of the susceptible rice variety Lijiang Xintuan Heigu was treated at a concentration of 1.0 mg / ml for 12 hours, and...

Embodiment 2

[0018] 10.0 mg / ml of fusion protein was used to treat rice root tissue for 12 hours, and the formation of callose was directly observed after aniline blue staining.

[0019] Centrifuge the expression product of the effector protein gene of Magnaporthe grisea to collect the cells, and suspend the cells with freshly prepared buffer (20mM Tris-HCl, pH7.4; 200mM NaCl; 1mM EDTA; 10mM β-mercaptoethanol) . The supernatant was collected by centrifugation after ultrasonography to disrupt the cells in an ice-water bath. The treatment is to treat the root tissue of the disease-resistant rice variety Nipponbare at a concentration of 10.0 mg / ml after purification of the prokaryotic expression product of the gene for 12 hours, and directly stain with aniline blue to observe callose. As a control, the prokaryotic expression product of the gene was purified, and the root tissue of the susceptible rice variety Lijiang Xintuan Heigu was treated at a concentration of 10.0 mg / ml for 12 hours, an...

Embodiment 3

[0023] 2.0 mg / ml fusion protein was used to treat rice root tissue for 12 hours, and the formation of callose was directly observed after aniline blue staining.

[0024] Centrifuge the expression product of the effector protein gene of Magnaporthe grisea to collect the cells, and suspend the cells with freshly prepared buffer (20mM Tris-HCl, pH7.4; 200mM NaCl; 1mM EDTA; 10mM β-mercaptoethanol) . The supernatant was collected by centrifugation after ultrasonography to disrupt the cells in an ice-water bath. The treatment is to treat the root tissue of the disease-resistant rice variety Nipponbare at a concentration of 2.11 mg / ml after purification of the prokaryotic expression product of the gene for 12 hours, and directly stain with aniline blue to observe callose. As a control, after the prokaryotic expression product of the gene was purified, the root tissue of the susceptible rice variety Lijiang Xintuan Heigu was treated at a concentration of 2.11 mg / ml for 12 hours, and ...

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Abstract

The invention relates to a method for treating paddy rice tissues by using a prokaryotic expression product of a magnaporthe grisea effect protein gene and inducing generation of callose, belonging to the technical field of plant protection and biology. The method is an improvement on the prior art, comprises a prokaryotic expression technology which is the same as that of the conventional method, a protein purifying technology, the wavelength ranges of exciting light and emitted light of a used fluorescence microscope and an aniline blue dyeing method, and is characterized by comprising the following steps of: treating the root tissues of Nipponbare serving as a disease-resistant paddy rice variety with the prokaryotic expression product which is 1.0-10.0 mg/ml in concentration of the magnaporthe grisea effect protein gene is used for treating the root tissues for 12 hours; performing aniline blue dyeing; and directly observing the formation of callose under the fluorescence microscope. The method is easy, convenience, accurate and rapid; and a large amount of spontaneous fluorescence interference can be avoided effectively, and the situation of basic defense reaction of paddy rice can be obtained directly and qualitatively.

Description

Technical field: [0001] The invention relates to a method for treating rice root tissue with a prokaryotic expression product to induce callose production, in particular to a method for treating rice root tissue with a prokaryotic expression product of the rice blast fungus effector protein gene to induce callose production, belonging to plants fields of conservation and biotechnology. Background technique: [0002] Plants and their pathogens have co-evolved for millions of years, and natural selection has prompted plants to produce a variety of recognition and resistance mechanisms to prevent and limit pathogen infection. The evolution of plants also promotes the evolution of pathogenic bacteria genes to adapt or resist plant defense responses to infect hosts to achieve the ultimate goal of pathogenicity. As a result of host-pathogen evolution, the genes involved in the evolution of pathogens diversify, especially the genes encoding pathogen effector proteins. Although ef...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/04C07K14/37
Inventor 杨静李成云刘林朱有勇王凯施竹凤
Owner YUNNAN AGRICULTURAL UNIVERSITY
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