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Primers, kits and methods for triple fluorescent quantitative PCR detection of dtmuv, edsv and h9 subtype aiv

A technology for detection kits and detection primers, which are applied in the directions of microorganism-based methods, biochemical equipment and methods, and the determination/inspection of microorganisms to achieve the effect of good repeatability and good amplification effect.

Active Publication Date: 2017-11-14
INST OF ANIMAL HEALTH GUANGDONG ACADEMY OF AGRI SCI
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  • Abstract
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Problems solved by technology

So far, no kits have been reported for simultaneous detection of duck Tembusu virus, egg drop syndrome virus and H9 subtype avian influenza virus

Method used

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  • Primers, kits and methods for triple fluorescent quantitative PCR detection of dtmuv, edsv and h9 subtype aiv
  • Primers, kits and methods for triple fluorescent quantitative PCR detection of dtmuv, edsv and h9 subtype aiv
  • Primers, kits and methods for triple fluorescent quantitative PCR detection of dtmuv, edsv and h9 subtype aiv

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Embodiment Construction

[0067] The present invention will be further described below in conjunction with the examples, but not limited thereto.

[0068] 1 Materials and methods

[0069] 1.1 Viruses Duck Tembusu virus JM strain, egg drop syndrome virus and H9 subtype avian influenza virus are preserved by our laboratory.

[0070] 1.2 Reagents One Step SYBR PrimeScript RT-PCR Kit II, DNA Molecular Weight Standards, pMD18-T Vector, Takara MiniBEST DNA / RNA Extraction Kit, Gel Recovery Kit were purchased from Dalian Bao Biological Engineering Co., Ltd. DH5α Competent was purchased from Tiangen company, and other reagents were of analytical grade.

[0071] 1.3 Primer design and synthesis Download the complete gene sequence of Duck Tembusu virus, Egg drop syndrome virus and H9 subtype avian influenza virus gene sequence from GenBank, and use Oligo 6.0 to design specific primers for their conserved regions. The primers were synthesized by Invitrogen, and the sequences are shown in Table 1.

[0072] Table ...

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Abstract

The invention discloses a triple fluorescent quantitative PCR detection primer, kit and method for duck Tembusu virus, egg drop syndrome virus and H9 subtype avian influenza virus. The detection primers for duck Tembusu virus, egg drop syndrome virus and H9 subtype avian influenza virus are respectively shown in SEQ ID NO.1-6. The detection primer and kit of the present invention can detect EDSV, H9 AIV and DTMUV simultaneously, and do not interact with duck plague virus, goose parvovirus, muscovy duck parvovirus, Escherichia coli genomic DNA, duck hepatitis A virus type 1, and duck hepatitis A virus type 3 , Muscovy duck reovirus and Newcastle disease virus RNA cross-reacts, and the detection limits for EDSV, H9 AIV and DTMUV nucleic acids are 8.0, 4.8 and 1.3 copies respectively, which can be used for the qualitative and quantitative analysis of EDSV, H9 AIV and DTMUV Quantitative detection.

Description

technical field [0001] The invention belongs to the field of molecular detection, and in particular relates to a triple fluorescent quantitative PCR detection primer, kit and method for duck Tembusu virus, egg drop syndrome virus and H9 subtype avian influenza virus. Background technique [0002] Duck Tembusu Virus Disease is an acute infectious disease caused by Duck Tembusu Virus (DTMUV). The disease can cause laying ducks and breeding geese to show obvious symptoms of egg production decline in a short period of time, and the course of the disease can last for several weeks. Recently, there have also been reports of infection in laying hens and meat ducks. The causative agent of the disease belongs to the family Flaviviridae, the genus Flavivirus, and Tembusu virus. Although the mortality rate of the disease is not high, the course of the disease is long and the potential harm is huge. The duck Tembusu virus genome is about 11 kb long, and the sequence is 5'-C-prM-E-NS1...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/93
Inventor 孙敏华董嘉文李林林袁建丰邝瑞欢张建峰刘志成张春红
Owner INST OF ANIMAL HEALTH GUANGDONG ACADEMY OF AGRI SCI
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