Test strip for quickly detecting cucumber green mottle mosaic virus (CGMMV)

A technology of green mottled flowers and test strips, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of high detection cost, long detection time, and inability to detect on-site, and achieve short detection time, low cost, and improved sensitivity and accuracy effects

Active Publication Date: 2014-01-01
POMOLOGY RES INST FUJIAN ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to solve the above-mentioned shortcomings such as long detection time, inability to detect on-site, high detection cost, etc., to provide a specificity, high sensitivity, simple and fast, low cost, suitable for cucumber green mottled flowers and leaves that can be detected on-site Colloidal gold immunochromatographic test strip for virus (CGMMV) and preparation method thereof

Method used

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  • Test strip for quickly detecting cucumber green mottle mosaic virus (CGMMV)
  • Test strip for quickly detecting cucumber green mottle mosaic virus (CGMMV)
  • Test strip for quickly detecting cucumber green mottle mosaic virus (CGMMV)

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Effect test

Embodiment 1

[0028] The preparation method of detecting cucumber green mottled mosaic virus immune colloidal gold test strip of the present invention, such as figure 1 , 2 shown, including the following steps:

[0029] (1) Preparation of CGMMV polyclonal antibody: Amplification of coat protein of cucumber green mottle mosaic virus Guangxi isolate by RT-PCR method cp gene and cloned into the pMD-18T vector. Will cp The gene was inserted into the prokaryotic expression vector pET-32a (+), and the recombinant plasmid CGMMVpET-32aCP was constructed, and its sequence was confirmed. Then the recombinant plasmid of CGMMVpET-32aCP was transformed into Escherichia coli BL21 (DE3) pLysS strain. Induce for 25 hours at a final concentration of IPTG of 0.1 mmol / L, a constant temperature of 23 °C, and a shaking speed of 90 rpm to obtain soluble expression of the foreign fusion protein of CGMMV-CP, and purify the expression supernatant with a Ni Sepharose FF column , the target protein was centrif...

Embodiment 2

[0037] Using the present invention to detect cucumber green mottle mosaic virus immune colloidal gold test strips to detect cucumber green mottle mosaic virus method, comprising the following steps:

[0038] (1) Collection and processing of samples to be tested: Weigh 0.5 g of Cucurbitaceae plant leaves (or roots, stems) into a 2 mL sample tube, add 0.01 mol / L pH7.4 phosphate buffer, mash with a glass rod, and then Centrifuge at 8000 rpm for 5 minutes, take the supernatant, which is the sample solution to be tested;

[0039] (2) Before the test, the colloidal gold test strips for the detection of cucumber green mottle mosaic virus immune colloidal gold were returned to room temperature and then opened for use;

[0040] (3) Take the colloidal gold test strip for the detection of cucumber green mottle mosaic virus immunity from the aluminum foil bag, and drop the sample solution of step (1) on the sample pad of the test strip, or insert the test strip into the step ( 1) The liq...

Embodiment 3

[0043] specificity test

[0044] Use the invention to detect cucumber green mosaic mosaic virus, watermelon mosaic virus, zucchini yellow mosaic virus, tobacco mosaic virus, cucumber mosaic virus virus) and pumpkin mosaic virus (Squash mosaic virus) positive samples, the specificity of this test paper has been proved, including the following steps:

[0045] (1) Collection and processing of samples to be tested: Weigh 0.5 g of the above-identified virus-positive plant leaves into a 2 mL sample tube, add 0.01 mol / L pH7.4 phosphate buffer, mash with a glass rod, and then centrifuge at 8000 rpm After 5 minutes, take the supernatant, which is the sample solution to be tested;

[0046] (2) Before the test, the colloidal gold test strips for the detection of cucumber green mottle mosaic virus immune colloidal gold were returned to room temperature and then opened for use;

[0047] (3) Take the colloidal gold test strip for the detection of cucumber green mottle mosaic virus immunit...

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Abstract

The invention discloses an immune colloidal gold test strip for quickly detecting a cucumber green mottle mosaic virus (CGMMV) and a preparation method of the immune colloidal gold test strip. The preparation method comprises the following steps: dripping a purified CGMMV polyclonal antibody solution on a detection line of a nitrocellulose membrane; dripping a goat anti-rabbit polyclonal antibody on a quality control line of the nitrocellulose membrane; sequentially attaching a sample pad made of water-absorbing filter paper, a CGMMV immune colloidal gold pad, the nitrocellulose membrane and water-absorbing paper to a polyvinyl chloride (PVC) adhesive plate with an adhesive sticker so as to prepare the test strip. The test strip can be used for detecting the CGMMV in a cucurbitaceous plant and is applicable to detection and diagnosis of a viral disease caused by the CGMMV. The method is simple and quick. The test strip is low in cost. A result can be obtained within 5 to 10 minutes and can be visually and directly observed by naked eyes. The test strip is applicable to quick and extensive screening and detection of diseases through basic agricultural departments at all levels, farmers and entry-exit inspection departments, facilitates timely and effective control and elimination of an epidemic situation, and has a wide application prospect.

Description

technical field [0001] The invention relates to a test strip for detecting cucumber green mottle mosaic virus in plants and a preparation method, in particular to an immune colloidal gold test strip for detecting cucumber green mottle mosaic virus in Cucurbitaceae and a preparation method. Background technique [0002] Cucumber green mottle mosaic virus ( Cucumber green mottle mosaic virus , CGMMV) belonging to the family Turnipmoviridae ( Tymoviridae ) Tobacco mosaic virus ( Tobamovirus ), is one of the important viruses of Cucurbitaceae, mainly infecting cucumbers, watermelons, melons, gourds, pumpkins and other Cucurbitaceae plants. The virus was first discovered on cucumbers in the United Kingdom in 1935, and subsequently occurred in Greece, Israel, France, India, Japan, Korea, Indonesia and Pakistan, causing serious economic losses to the production of cucurbit crops. CGMMV occurred relatively late in my country. In 1987, the virus was isolated and identified from ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569G01N33/531
CPCG01N33/531G01N33/56983
Inventor 范国成林雄杰林志铿胡菡青陈庆河翁启勇
Owner POMOLOGY RES INST FUJIAN ACAD OF AGRI SCI
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