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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: 2011-08-31
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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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

Examples

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Effect test

Embodiment 1

[0023] Example 1. Optimization of RT-LAMP reaction conditions for rice black-streaked dwarf virus

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

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

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 prediction, belonging to the field of agricultural science and technology. Background technique [0002] Rice black-streaked dwarf virus (RBSDV) is a serious viral disease on rice, belonging to the genus Fijivirus of the plant Reoviridae, with spherical virions, mainly composed of The plant flies are spread by the planthopper. The early symptoms in the field are mainly 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 are formed, which has a great impact on the yield of rice, and even in severely diseased fields. Can lead to no production. In addition to infecting rice, the virus can also infect maize (maize rough dwarf disease), wheat (wheat green dwarf) and various grass weeds. In 1963 and 1966, it bec...

Claims

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Application Information

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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 ACADEMY OF AGRICULTURAL SCIENCES
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