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Primer for detecting porcine sapelo virus, Taqman-MGB probe and kit

A pig sape and kit technology, which is applied in the field of detection of animal pathogenic microorganisms, can solve problems such as inability to quantitatively analyze, long technical cycle, and insufficient sensitivity, and achieve the effects of simplified optimization steps, high sensitivity, and accurate experimental results

Active Publication Date: 2013-07-17
WENS FOOD GRP CO LTD
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

[0005] At present, the conventional methods for identifying porcine Sapero virus include virus isolation, virus neutralization test, lesion morphology observation, antigen analysis, RT-PCR, etc. Due to the long cycle of these technologies, the operation is cumbersome or only qualitative detection can not be quantitatively analyzed , and sometimes the sensitivity is not enough, especially for samples with low virus content, so it is limited in practical application
There is no report on the detection of this virus using Taqman-MGB real-time fluorescent quantitative RT-PCR

Method used

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  • Primer for detecting porcine sapelo virus, Taqman-MGB probe and kit
  • Primer for detecting porcine sapelo virus, Taqman-MGB probe and kit
  • Primer for detecting porcine sapelo virus, Taqman-MGB probe and kit

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Embodiment 1

[0029] 1) Design primers and TaqMan-MGB probes

[0030] The whole genome sequence of porcine sapero virus was retrieved from Genbank (the reference strain Genbank accession numbers are JX286666.1, NC_003987.1 and HQ875059.1), and the sequences were compared by DNAstar software to find out the porcine sapero virus There are 10 conserved sequences specific to the viral genome, which were logged at http: / / www.ncbi.nlm.nih.gov / and BLASTed to determine their conservation. Using Oligo 6.0 software to analyze the base composition and stability of these conserved sequences, and comprehensively consider the principles of primer and probe design, and finally screen out the most suitable conserved sequence, which is located in the PSV genome The 153bp to 260bp of the 5'untranslated region (strain Genbank accession number JX286666.1).

[0031] Use professional primer design software to design primers for this conserved sequence, and obtain multiple pairs of specific primers and probe se...

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Abstract

The invention discloses a primer for detecting porcine sapelo virus, a Taqman-MGB probe and a kit, and belongs to the technical field of animal pathogenic microorganism detection. A nucleotide sequence of the primer for detecting the porcine sapelo virus is represented by SEQ ID NO:1-2; and a DNA sequence of the Taqman-MGB probe is represented by SEQ ID NO:3. The primer and the Taqman-MGB probe are high in specificity, high in stability and high in detection accuracy; the Taqman-MGB diagnosis kit has the characteristics of high detection speed, high sensitivity, high stability, simplicity in quantification and low false positive and is favorable for large-scale and automatic detection analysis. By adopting a real-time fluorescent quantitation RT-PCR method for the Taqman-MGB, the shortcomings of complicated virus separation operation process, difficulty in authentication, low detection speed of the conventional PCR, simplicity in pollution, need of electrophoresis caused by amplification and small number of samples to be detected at each time are overcome.

Description

technical field [0001] The invention relates to the technical field of detection of animal pathogenic microorganisms, in particular to a primer, a Taqman-MGB probe and a kit for detecting porcine Sapero virus. Background technique [0002] Porcine sapelovirus (PSV) is a single-stranded positive-sense RNA virus belonging to the family Picornaviridae and the genus Enterovirus. Porcine sapero virus was previously classified as porcine enterovirus A, and was once named PEV-8, but Krumbholz et al published a paper entitled "Sequencing of Porcine Enterovirus Groups II and III Reveals Unique Features of Both Virus Groups", which can distinguish it from other porcine enteroviruses based on its L and 2A genes, so it is classified as a new genus. Taxonomy of Viruses - Eighth Report of the International Committee on Taxonomy of Viruses (ICTV) finally named PEV-8 the genus Saperovirus, which includes porcine Saperoviruses, avian Saperoviruses, and simian Saperoviruses. [0003] Porci...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/93
Inventor 陈俊伟
Owner WENS FOOD GRP CO LTD
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