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Reverse transcription loop-mediated isothermal amplification (RT-LAMP) method for rapidly detecting rice black-streaked dwarf viruses in plant hopper

A technology of RT-LAMP, rice black stripe dwarfing, applied in the field of agricultural science, can solve the problems of high sequence homology and difficult to distinguish

Inactive Publication Date: 2013-01-09
JIANGSU ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Since these two viruses have very similar characteristics in many aspects, traditional plant virus detection methods, such as electron microscope observation (similar in size and shape of virus particles), traditional biological inoculation identification (similar in symptoms, mediators, and host ranges) ), serological detection (sequences still have high homology) and other methods are difficult to distinguish

Method used

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  • Reverse transcription loop-mediated isothermal amplification (RT-LAMP) method for rapidly detecting rice black-streaked dwarf viruses in plant hopper
  • Reverse transcription loop-mediated isothermal amplification (RT-LAMP) method for rapidly detecting rice black-streaked dwarf viruses in plant hopper

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Effect test

Embodiment 1

[0023] Embodiment 1, optimization of rice black-streaked dwarf virus RT-LAMP reaction conditions

[0024] ①Design of rice black-streaked dwarf virus RT-LAMP primers.

[0025] According to the reported rice black-streaked dwarf virus (AF227206.1, http: / / www.ncbi.nlm.nih.gov / nuccore / 183229399; EU784843.1, http: / / www.ncbi.nlm.nih .gov / nuccore / 209867306) S10 nucleotide sequence, find out the relatively conserved region 950-1467bp of the virus nucleic acid after analysis by software DNAstar and design primers. Using a relatively conserved sequence (950-1467 bp) of the virus as a template, primers were designed using the online LAMP primer design software primerExplorer V4 (http: / / primerexplorer.jp / elamp4.0.0 / index.html). After uploading the template sequence, the system software calculates and obtains the preliminary LAMP primer combination (F3, B3, FIP, BIP), and then provides parameters such as the stability of the 3' end or 5' end of the primer and the primer dimer through the ...

Embodiment 2

[0032] Embodiment 2, the specificity test of rice black-streaked dwarf virus RT-LAMP reaction

[0033] In order to verify the specificity of the RT-LAMP method, the southern rice black-streaked dwarf virus (SRBSDV) of the same genus as RBSDV was selected as a control. Using the total RNA of SBPH carrying RBSDV and SBPH carrying SRBSDV as templates, the reaction was carried out according to the RT-LAMP reaction system in Example 1. The results showed that the total RNA templates of SBPH carrying SRBSDV and avirulent SBPH had no amplified products; while SBPH carrying RBSDV could amplify the target product ( figure 2 ). This indicates that the specificity of the RT-LAMP primer is better.

[0034] The invention discloses a method for quickly detecting rice black-streaked dwarf virus in planthoppers, through which it is possible to quickly diagnose whether planthoppers carry rice black-streaked dwarf virus.

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Abstract

The invention provides a rapid detection technology for rice black-streaked dwarf viruses in plant hopper and a method for applying the rapid detection technology to disease diagnosis and prediction. By the method, the interference of southern rice black-streaked dwarf viruses can be eliminated, and whether the rice black-streaked dwarf viruses are carried in the plant hopper is rapidly identified, so that a pertinent control strategy is adopted to reduce the loss caused by diseases.

Description

technical field [0001] The invention relates to a rapid detection technology of rice black-streaked dwarf virus in planthoppers and its application in disease diagnosis and forecasting, belonging to the field of agricultural science and technology. Background technique [0002] Rice black-streaked dwarf virus (RBSDV) is a serious virus disease on rice, belonging to the plant Reoviridae Fijivirus genus (Fijivirus), the virion is spherical, mainly composed of SBPH spreads. In the early stage of field symptoms, the main symptoms are dwarf plants and dark green leaves. In the later stage, wax drop-like protrusions appear on the leaves, leaf sheaths and stems, and then black stripes form. This has a great impact on the yield of rice. Severely diseased fields even May result in no yield. In addition to infecting rice, the virus can also infect corn (maize rough dwarf disease), wheat (wheat green dwarf) and various grass weeds. It was popular in Jiangsu, Shanghai and Zhejiang in ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11
Inventor 周彤周益军杜琳琳兰莹徐秋芳孙枫范永坚
Owner JIANGSU ACAD OF AGRI SCI
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