Method for rapidly detecting Chinese tomato yellow leaf curl virus (TYLCCNV) by using LAMP (Loop-mediated isothermal amplification)

A technology for tomato yellowing leaf curl and virus, which is applied in the directions of biochemical equipment and methods, microbial determination/inspection, etc., to achieve the effects of low cost, strong specificity and high sensitivity

Inactive Publication Date: 2014-05-28
SOUTHWEST UNIVERSITY
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Problems solved by technology

At present, there are few studies on the application of this technology to the detection of plant viruses, and there is no report on the detection of TYLCCNV using this technology.

Method used

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  • Method for rapidly detecting Chinese tomato yellow leaf curl virus (TYLCCNV) by using LAMP (Loop-mediated isothermal amplification)
  • Method for rapidly detecting Chinese tomato yellow leaf curl virus (TYLCCNV) by using LAMP (Loop-mediated isothermal amplification)
  • Method for rapidly detecting Chinese tomato yellow leaf curl virus (TYLCCNV) by using LAMP (Loop-mediated isothermal amplification)

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Embodiment Construction

[0022] The present invention will be further described below in conjunction with specific embodiments:

[0023] Take tobacco plant leaves infected with common tobacco leaf curl pathogens such as TYLCCNV, TbLCYNV, AYVV, TbCSV, PaLCuCNV, and implement the following detection according to the method of the present invention. The results are as follows image 3 Shown. From this figure, it can be clearly determined that only LAMP-characteristic diffuse nucleic acid bands can be detected in plants infected with TYLCCNV, which confirms the feasibility and accuracy of the method of the present invention for detecting TYLCCNV. The specific operations are as follows:

[0024] (1) Use CTAB method to extract total DNA from diseased leaves of plants:

[0025] A. Take 0.2g of diseased leaves of the plant, grind thoroughly with liquid nitrogen, add to a 1.5mL centrifuge tube pre-placed with 1mL CTAB extract, vortex and mix for 30s, and bath at 65℃ for 30min;

[0026] B. Add an equal volume of pheno...

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Abstract

The invention discloses a method for rapidly detecting Chinese tomato yellow leaf curl virus (TYLCCNV) by using a loop-mediated isothermal amplification (LAMP) technology. The method comprises a reaction system and a specific primer group contained in the reaction system. The method comprises the steps of extracting DNA (Deoxyribonucleic Acid), carrying out LAMP, carrying out electrophoresis detection or fluorescent staining and the like. The method has the advantages of strong specificity, high sensitivity, simplicity, convenience, rapidness, low cost and the like, and special instruments such as a PCR (Polymerase Chain Reaction) instrument and equipment such as gel electrophoresis equipment are not required, so that the rapid detection on virus disease samples of laboratories and fields is facilitated.

Description

technical field [0001] The invention relates to the technical field of plant disease detection, in particular to a loop-mediated isothermal amplification (Reverse Transcription Loop-Mediated Isothermal Amplification, LAMP) molecular detection method, in particular to a method for rapidly detecting Chinese tomato yellow leaf curl virus using LAMP . Background technique [0002] Chinese Tomato Yellow Leaf Curl Virus (TYLCCNV) is a Geminiviridae ( Geminivirade ) Begomovirus ( Begomovirus ) virus. Since TYLCCNV has many types of hosts, it can infect tobacco, tomato, pumpkin, cassava, cotton and other crops, and it occurs in a wide range, such as Sichuan, Yunnan, Guangxi, Guangdong and Fujian. Early monitoring of viral diseases in the field is of great significance. Currently, the methods for detecting TYLCCNV are mainly serological methods and PCR techniques, both of which have the characteristics of fast speed, high sensitivity and good specificity. However, the disadvant...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68
CPCC12Q1/6844C12Q1/70C12Q2531/119
Inventor 王春艳青玲李科李茵
Owner SOUTHWEST UNIVERSITY
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