Detection kit for detecting bovine coronavirus (BCV) and application method thereof

A detection kit and coronavirus technology, applied in the detection kit and application field of bovine coronavirus BCV, can solve the problems of non-specific product amplification in complex systems, insufficient amplification efficiency, and difficulty in popularization and promotion, and achieve Effects of reducing non-specific amplification reactions, improving amplification efficiency and sensitivity, and reducing residence time

Pending Publication Date: 2020-04-24
QINGHAI ACAD OF ANIMAL SCI & VETERINARY MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Molecular biology detection methods mainly include RT-PCR, nested PCR, double PCR and real-time fluorescent quantitative PCR methods, etc., but they also have their limitations. For the amplification of complex systems, there will be non-specific product amplification and the amplification efficiency is not high enough. , The detection limit is not low enough, real-time quantitative PCR also needs expensive instruments, it is difficult to popularize in the grassroots laboratories
These shortcomings limit the application of the above methods in clinical diagnosis

Method used

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  • Detection kit for detecting bovine coronavirus (BCV) and application method thereof
  • Detection kit for detecting bovine coronavirus (BCV) and application method thereof
  • Detection kit for detecting bovine coronavirus (BCV) and application method thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1: the establishment of nano-PCR method

[0029] 1. Primer design:

[0030] Referring to the analysis of the genome sequence of BCV (accession number: MH475915) published in GenBank, the Primer3.0 software was used to design and amplify the full-length sequence primers of the BCV-N gene, which were screened after comparison with the published full-length sequence of the N gene. Primers were designed to amplify the 396bp fragment in the conserved region. Primers were synthesized by Suzhou Jinweizhi Biotechnology Co., Ltd.:

[0031]

[0032]

[0033] 2. Preparation of standard products:

[0034] After the BCV virus proliferated in Vero cells, the RNA was extracted, and after reverse transcription, the full-length sequence of the N gene was amplified with P1 / P2 primers as a template, and the size of the obtained product was 1344bp. After gel recovery and purification, it was ligated into the pET-28a vector to construct a recombinant plasmid, and after the...

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Abstract

The invention discloses a detection kit for detecting BCV and a method for applying the same to non-disease detection. The detection kit for detecting the BCV comprises an amplification primer as shown in SEQ ID NO. 3 and an amplification primer as shown in SEQ ID NO. 4. The detection kit for detecting the BCV and the application method thereof can rapidly and specifically detect the BCV.

Description

technical field [0001] The present invention relates to a test kit for detecting a specific virus and a method for using the test kit in non-disease detection purposes. Specifically, the present invention relates to a detection kit for detecting bovine coronavirus BCV and a method for using the non-disease detection . Background technique [0002] Bovine Coronavirus (BCV) is a non-segmented single-stranded positive-sense RNA virus with an envelope. Virus particles are pleomorphic, but basically spherical, with a diameter of 65-210nm and a genome size of 31043bp. They mainly encode five structural proteins, including nucleocapsid protein (N, 52kD) from the 5' end to the 3' end, Membrane protein (M, 25kD), small capsid protein (E, 8kD), spike protein (S, 180kD) and hemagglutinin esterase protein (HE, 65kD). Among them, the N gene is highly conserved and is often used for nucleic acid detection and identification of BCV; although the homology of the M protein gene sequence is...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/686C12R1/93
CPCC12Q1/701C12Q1/686C12Q2565/125
Inventor 郭志宏张学勇朵红付永沈秀英马怡隽
Owner QINGHAI ACAD OF ANIMAL SCI & VETERINARY MEDICINE
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