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PCR-RFLP primer and method for rapidly identifying genotypes of duck circoviruses

A PCR-RFLP and duck circovirus technology, which is applied in the field of PCR-RFLP primers for rapid identification of duck circovirus genotypes, can solve the problems of cumbersome optimization of double PCR conditions, achieve high speed, fill research gaps, and fast detection accurate effect

Inactive Publication Date: 2015-12-23
INST OF ANIMAL HUSBANDRY & VETERINARY FUJIAN ACADEMY OF AGRI SCI
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Problems solved by technology

At present, there have been research reports on the multiplex PCR method for the rapid identification of duck circovirus genotypes (genotype 1 and genotype 2), but this method needs to target duck circovirus genotypes (genotype 1 and genotype 2) respectively. ) to design specific primers and establish double PCR detection, but the optimization of double PCR conditions is cumbersome

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  • PCR-RFLP primer and method for rapidly identifying genotypes of duck circoviruses

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Embodiment 1

[0022] 1. Virus strains:

[0023] Duck circovirus genotype 1 (representative strain GenBank accession number EF451157) and duck circovirus genotype 2 (representative strain GenBank accession number GQ423745) were both preserved by the Institute of Animal Husbandry and Veterinary Medicine, Fujian Academy of Agricultural Sciences.

[0024] 2. Primer design and synthesis

[0025] Primers P1 and P2 were designed according to the genetic characteristics of the gene coding region of the replication protein gene of duck circovirus (genotype 1 and genotype 2), wherein the sequences of primers P1 and P2 are: P1: 5'-TGCGCCAAAGAGTCGACATAC-3', downstream primer P2 : 5'-CAAATTGGTCTCAGTAGTTTATT-3'.

[0026] 3. PCR amplification

[0027] The suspected disease materials submitted for clinical inspection were operated according to the instructions of TransDirectAnimalTissuePCRKit (Beijing Quanshijin Biotechnology Co., Ltd.). Take 40μL of AD1Buffer and add 10μL of AD2Buffer. To detect the di...

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Abstract

The invention discloses a PCR-RFLP primer and method for rapidly identifying the genotypes of duck circoviruses. The primer comprises an upstream primer body P1:5'-TGCGCCAAAGAGTCGACATAC-3' and a downstream primer body P2:5'-CAAATTGGTCTCAGTAGTTTATT-3'. According to the method, distinguishing is carried out through the difference of restriction enzyme cutting sites contained in coding regions of copied protein genes of the duck circoviruses in different genotypes (the first genotype and the second genotype). The method includes the steps that DNA is extracted, Kpn I restriction enzyme cutting is carried out after PCR amplification is carried out to obtain corresponding target fragments, and then RFLP analysis is carried out. The identification method is rapid and easy to operate and high in accuracy.

Description

Technical field [0001] The present invention is the field of rapid diagnosis of veterinary infectious diseases, and it involves a PCR-RFLP primer and method that quickly identifies duck ring virus genotypes. Background technique [0002] Since Hattermann's first reported DuckCircovirus (DUCV) et al., Scholars at home and abroad have successively studied duck ring virus infection, but currently reports differently in infection.Fringuelli et al. Fa's positive rates detected from Hungary's illness or death duck were 84%(85 / 101), which speculated that the virus may be popular in the local area;The tests collected during the year showed that the detection rate of ring virus was 38.2%(13 / 34), and the study of Banda and others showed that the detection rate of the duck group raised in the Long Island area of New York was very low; zhan, etc.A survey of duck ring virus in Shandong Province, some of the cherry valley duck groups, which occurred in Shandong Province, found that the positiv...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/93
CPCC12Q1/701C12Q1/6858
Inventor 万春和黄瑜陈红梅陈翠腾程龙飞傅光华傅秋玲施少华
Owner INST OF ANIMAL HUSBANDRY & VETERINARY FUJIAN ACADEMY OF AGRI SCI
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