Real-time fluorescent RT-PCR method for detecting whether arabis mosaic virus, bean yellow mosaic virus and lily symptomless virus infect host

A lily asymptomatic virus, RT-PCR technology, applied in biochemical equipment and methods, fluorescence/phosphorescence, microbial determination/inspection, etc., can solve the problems of reduced quality of cut flowers, low sensitivity, low concentration of seed bulb virus, etc. Achieve high sensitivity and specificity, and simple detection procedures

Inactive Publication Date: 2010-10-27
SHENZHEN AUDAQUE DATA TECH
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AI Technical Summary

Problems solved by technology

The growth potential of gladiolus seedlings infected by BYMV was weakened, white spots appeared on the leaves, and the flowers were variegated, resulting in a decrease in the quality of cut flowers and a decrease in bulb yield. Morbidity can be as high as 67% to 100%, resulting in 59% to 96% yield loss
The infection rate of lily by LSV is as high as 73%. When it is infected alone, it has no obvious symptoms. Under natural conditions, it is often co-infected with other viruses, resulting in serious mosaic leaves and dwarfing of the whole plant, affecting the yield of lily. and business value
[0003] The detection of Arabidopsis mosaic virus, lily asymptomatic virus and bean yellow mosaic virus can be detected by conventional enzyme-linked immunosorbent assay (ELISA), but this detection method has low sensitivity and bias; especially when gladiola and lil

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Detection of host A (gladiolus) infected by Arabidopsis thaliana mosaic virus, bean yellow mosaic virus and lily asymptomatic virus by multiplex RT-Real time PCR

[0051] Host A (gladiolus) is detected, specifically comprising the following steps:

[0052] Step a, extracting total RNA from host A (gladiolus); removing substances such as polysaccharides and phenols that affect the RT-Real time PCR reaction;

[0053] Step b, dilute the total RNA to 10 -2 times RNA solution;

[0054] Step c, using the RNA solution as a template to perform a real-time RT-Real time PCR reaction;

[0055] Real time RT-PCR reaction system: the total volume is 10 μL reaction system. 10×One Step RNA PCR Buffer 1μL, 25mmol / L MgCl 2 2μL, 10mmol / L dNTP Mixture 1μL, 40U / μL RNase Inhibitor 0.2μL, 5 U AMV Reverse Transcriptase XL 0.2μL, 5U AMV-Optimized Taq 0.2μL, 10μmol / L primer ArMV, primer BYMV and primer LSV upstream and downstream 0.4 μL for each primer, 0.4 μL for each of 5 μmol / L probe Ar...

Embodiment 2

[0074] Detection of host B (lily oriental) infected by Arabidopsis thaliana mosaic virus, bean yellow mosaic virus and lily asymptomatic virus by multiplex RT-Real time PCR

[0075] This detection step is basically the same as the detection method in Example 1, except that the hosts in step a are different.

[0076] Finally, the blank control and the negative control display all show negative, the positive control is positive, and the detection result of the sample to be tested shows positive. The result shows that the host B (Oriental lily) has been infected by Arabidopsis thaliana mosaic virus, bean yellow mosaic virus or lily. virus infection.

Embodiment 3

[0081] Detection of host C (Asian lily) infected by Arabidopsis thaliana mosaic virus, bean yellow mosaic virus and lily asymptomatic virus by multiplex RT-Real time PCR

[0082] This detection step is basically the same as the detection method in Example 1, except that the hosts in step a are different.

[0083] Finally, the blank control and the negative control display all show negative, the positive control is positive, and the detection result of the sample to be tested is positive. The result shows that the host C (Asian lily) has been infected by Arabidopsis thaliana mosaic virus, bean yellow mosaic virus or lily-free virus. virus infection.

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Abstract

The invention discloses a real-time fluorescent RT-PCR method for detecting whether arabis mosaic virus, bean yellow mosaic virus and lily symptomless virus infect a host. In the invention, according to the RNAs of the arabis mosaic virus, the bean yellow mosaic virus and the lily symptomless virus, a specific amplimer and a fluorescent probe are designed, RT-Realt time PCR reaction is carried out the RNA template of virus to be identified by means of the specific amplimer and the fluorescent probe, and whether the arabis mosaic virus, the bean yellow the mosaic virus and the lily symptomless virus infect the host is rapidly detected by carrying out real-time fluorescent detection on RT-Real time PCR products.

Description

technical field [0001] The invention relates to a virus species detection technology, in particular to a rapid detection technology for whether Arabidopsis thaliana mosaic virus, bean yellow mosaic virus and lily asymptomatic virus infect hosts. Background technique [0002] Arabis mosaic virus (ArMV), Bean yellow mosaic virus (BYMV) and Lily symptomless virus (LSV) are the main pathogens infecting lily and gladiolus crops, causing greater economic losses to growers. Arabidopsis mosaic virus has a wide range of hosts and can infect about 174 genera and 215 species of plants. It naturally infects and harms a variety of flowers, melons, beans and fruit trees, causing mosaic, yellowing, chlorosis, dwarfing, and shrinking Even symptoms such as necrosis are the entry plant quarantine pests stipulated by our country. BYMV has a wide range of hosts, and the main hosts are ornamental plants such as gladiolus and leguminous plants. The growth potential of gladiolus seedlings infec...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68G01N21/64C12N15/11
Inventor 郑耘陈枝楠陈富华李一农杨伟东
Owner SHENZHEN AUDAQUE DATA TECH
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