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Dual fluorescent quantitative RT-PCR (Reverse Transcription-Polymerase Chain Reaction) detection kit for detecting PEDV-TGEV (Porcine Epidemic Diarrhea Virus-Transmissible Gastroenteritis Virus) nucleic acid

A PEDV-TGEV, dual fluorescence technology, applied in the field of inspection and quarantine, to achieve the effect of convenient operation, good specificity and storage resistance

Inactive Publication Date: 2018-12-28
ZHEJIANG FORESTRY UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Due to the rapid onset of this disease, timely diagnosis is an effective means of prevention and control of this disease. At present, there is no method that can differentially diagnose the two pathogens that cause porcine viral diarrhea at one time.

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  • Dual fluorescent quantitative RT-PCR (Reverse Transcription-Polymerase Chain Reaction) detection kit for detecting PEDV-TGEV (Porcine Epidemic Diarrhea Virus-Transmissible Gastroenteritis Virus) nucleic acid
  • Dual fluorescent quantitative RT-PCR (Reverse Transcription-Polymerase Chain Reaction) detection kit for detecting PEDV-TGEV (Porcine Epidemic Diarrhea Virus-Transmissible Gastroenteritis Virus) nucleic acid
  • Dual fluorescent quantitative RT-PCR (Reverse Transcription-Polymerase Chain Reaction) detection kit for detecting PEDV-TGEV (Porcine Epidemic Diarrhea Virus-Transmissible Gastroenteritis Virus) nucleic acid

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Embodiment Construction

[0040] Compared with the SYBGreenI dye method, the Taq man probe fluorescence quantitative technology has higher specificity and sensitivity, mainly using the specific binding of the probe. The basic principle is: in the PCR amplification process, the 5'- 3' exonuclease activity, after the fluorescent probe hybridizes with the target fragment, when the Taq enzyme moves to the probe binding position during extension and replication, its exonuclease activity cuts the luminescent group at the 5' end of the probe and releases the fluorescence , is detected by a fluorescent quantitative PCR instrument, and when the probe is not combined with the target fragment, the quenching group at the 3' end of the probe interacts with the luminescent group, and no fluorescent signal appears. Therefore, during the PCR reaction, there is a one-to-one relationship between the number of released fluorescent groups and the PCR product. The present invention utilizes two different luminescent groups...

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Abstract

The invention relates to the field of inspection and quarantine and aims to provide a dual fluorescent quantitative RT-PCR (Reverse Transcription-Polymerase Chain Reaction) detection kit for detectingPEDV-TGEV (Porcine Epidemic Diarrhea Virus-Transmissible Gastroenteritis Virus) nucleic acid. The kit comprises (1) an inverse transcription and fluorescent quantitative RT-PCR amplification mixed liquid, wherein the mixed liquid comprises an inverse transcription and fluorescent quantitative amplification primer and a probe and has sequence of SEQ ID NO: 1-6 as shown in the specification, and the inverse transcription primer is a 12 basic group random primer; (2) negative control; (3) positive control, wherein the positive control comprises positive control plasmid for the porcine epidemic diarrhea virus and the transmissible gastroenteritis virus. Compared with the prior art, the kit has the advantages of being high in flux, low in cost, good in specificity and high in sensitivity.

Description

technical field [0001] The invention provides a dual fluorescence quantitative RT-PCR differential detection kit for simultaneously detecting porcine epidemic diarrhea virus (PEDV) and porcine transmissible gastroenteritis virus (TGEV) nucleic acids, which can realize simultaneous detection of two kinds of porcine infectious gastroenteritis virus (TGEV) nucleic acids. The pathogen of viral diarrhea syndrome belongs to the field of inspection and quarantine. Background technique [0002] Porcine epidemic diarrhea virus (PEDV) and porcine transmissible gastroenteritis virus of swine (TGEV) are currently the most important pathogens that endanger the diarrhea of ​​lactating and weaned piglets in pig farms in my country. The morbidity and mortality are high, and the onset and transmission are rapid, and it is difficult to control, which brings great losses to the pig industry. Once infected with the disease, piglets often exhibit watery diarrhea and are often dehydrated until th...

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6851C12R1/93
CPCC12Q1/6851C12Q1/701C12Q2600/16C12Q2521/107C12Q2531/113C12Q2563/107
Inventor 王晓杜宋厚辉吴瑗周莹珊邵春艳章先程昌勇姜胜孙静周彬宋泉江
Owner ZHEJIANG FORESTRY UNIVERSITY