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Method for simultaneously detecting three viruses and two kinds of viruses in chrysanthemum by multiple RT-PCR

A RT-PCR, virus technology, applied in the field of garden plant molecular biology, can solve the problem of multiple virus complex infection, save detection time and reagents and other consumables, and achieve the effect of efficient detection

Pending Publication Date: 2021-08-20
BEIJING AGRO BIOTECH RES CENT
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

PCR technology can amplify the genetic material of viruses in a short period of time, and has the advantages of rapidity, accuracy, and high sensitivity, but it can only detect one virus at a time. The pathogen needs to be repeated multiple times for PCR detection

Method used

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  • Method for simultaneously detecting three viruses and two kinds of viruses in chrysanthemum by multiple RT-PCR
  • Method for simultaneously detecting three viruses and two kinds of viruses in chrysanthemum by multiple RT-PCR
  • Method for simultaneously detecting three viruses and two kinds of viruses in chrysanthemum by multiple RT-PCR

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Embodiment Construction

[0022] 1. Experimental materials

[0023] Samples of chrysanthemum virus disease were collected from chrysanthemum planting areas in Shunyi, Pinggu, Yanqing and other suburban counties, and 1 g of fresh leaves were quick-frozen under liquid nitrogen and stored in a -80°C refrigerator.

[0024] 2. Primer design

[0025] The gene sequences of chrysanthemum B virus, tomato infertility virus, tomato spotted wilt virus, chrysanthemum dwarf viroid and chrysanthemum chlorotic mottle viroid were downloaded from NCBI database, and the gene sequences were compared by using clustalX software. Select the conserved region and use Primer 5.0 software to design primers. The design process requires that the annealing temperature of each primer be 50-60°C, the GC content be 40-60%, and try to avoid dimer and hairpin loop structures between each primer. In order to ensure the effect of electrophoretic differentiation, the size of the amplified sequences of different viruses and viroids differs...

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Abstract

The invention discloses a method for simultaneously detecting three viruses and two kinds of viruses in chrysanthemum by multiple RT-PCR, which adopts the following primer pair combination: CSVd-F and CSVd-R, the nucleotide sequences of which are shown as SEQ ID NO: 1 and SEQ ID NO: 2; CChMVd-F and CChMVd-R, the nucleotide sequences of which are shown as SEQ ID NO: 3 and SEQ ID NO: 4; CVB-F and CVB-R, the nucleotide sequences of which are shown as SEQ ID NO: 5 and SEQ ID NO: 6; TAV-F and TAV-R, the nucleotide sequences of which are shown as SEQ ID NO: 7 and SEQ ID NO: 8; TSWV-F and TSWV-R, the nucleotide sequences of which are shown as SEQ ID NO: 9 and SEQ ID NO: 10. The method provided by the invention can be used for effectively detecting plant viruses.

Description

technical field [0001] The invention relates to the field of garden plant molecular biology, in particular to a multiplex RT-PCR method for simultaneously detecting three kinds of viruses and two kinds of viruses in chrysanthemums. Background technique [0002] There are more than 20 kinds of viruses that infect chrysanthemums worldwide, and 14 kinds of viruses have been reported in my country at present. There are many types of viruses, wide transmission routes, and plant "cancer", and there are no ideal control measures. Early quarantine, prevention and control of spread, and suppression of transmission media are more effective prevention and control methods. Rapid virus detection is a prerequisite for virus prevention and treatment. [0003] At present, the detection techniques of chrysanthemum virus mainly include biological detection method, enzyme-linked immunosorbent assay, nucleic acid molecular hybridization technology, PCR technology and so on. The biological det...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/686C12N15/11C12R1/94
CPCC12Q1/701C12Q1/686C12Q2600/16C12Q2537/143C12Q2565/125
Inventor 陈东亮王森黄丛林罗昌刘华程曦
Owner BEIJING AGRO BIOTECH RES CENT
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