Primer probe group, kit and method for detecting G II type norovirus

A primer probe and kit technology, applied in the field of norovirus detection, can solve the problem of single norovirus genotype, etc., and achieve the effects of high sensitivity, short time consumption, and easy operation.

Pending Publication Date: 2021-08-06
INST OF ENVIRONMENTAL MEDICINE & OCCUPATIONAL MEDICINE ACAD OF MILITARY MEDICINE ACAD OF MILITARY SCI
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  • Abstract
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  • Claims
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Problems solved by technology

[0004] But the norovirus genotype detected by the method in the prior art is single

Method used

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  • Primer probe group, kit and method for detecting G II type norovirus
  • Primer probe group, kit and method for detecting G II type norovirus
  • Primer probe group, kit and method for detecting G II type norovirus

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Embodiment 1

[0060] Selection of Fluorescent RPA Detection Targets

[0061] The present invention selects the 294bp conserved specific sequence (SEQ ID No.4) of GII.4 type norovirus, specifically as follows:

[0062] 5'-GCTGGGCGAGGCTGCACTCCACGGCCCGGCATTCTATAGCAAAA TTAGCAAATTAGTCATTGCAGAGTTGAAGGAAGGTGGCATGGATTTTTA CGTGCCCAGACAAGAGCCAATGTTCAGATGGATGAGATTCTCAGATCT GAGCACGTGGGAGGGCGATCGCAATCTGGCTCCCAGTTTTGTGAATGA AGATGGCGTCGAGTGACGCCAACCCATCTGATGGGTCCGCAGCCAACC TCGTCCCAGAGGTCAACAATGAGGTTATGGCTCTGGAGCCCGTTGTTGGTGCCGCCA-3'。

[0063] Using the conserved specific sequence as the detection target, Shanghai Sangon Bioengineering Co., Ltd. carried out whole gene synthesis, and ligated the target fragment to the pUC57 plasmid with a length of 2710bp to construct a norovirus plasmid standard.

[0064] Primer design and screening

[0065] The length of RPA primers is recommended to be between 30-36bp, the GC content is between 20-70%, the melting temperature of the primers is between 50-100°C, the maxi...

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Abstract

The invention provides a primer probe group, a kit and a method for detecting G II type norovirus, and belongs to the technical field of norovirus detection. The primer probe group comprises an upstream primer NVF6, a downstream primer NVR6 and a fluorescent probe NVP9, wherein the nucleotide sequence of the upstream primer NVF6 is shown as SEQ ID No. 1; the nucleotide sequence of the downstream primer NVR6 is shown as SEQ ID No. 2; and the nucleotide sequence of the fluorescent probe NVP9 is shown as SEQ ID No. 3. The primer probe group provided by the invention can be used for specifically detecting G II type norovirus including G II. 4, G II. 3, G II. 6 and G II. 17, and has no cross reaction with astrovirus, MS2 bacteriophage, human rotavirus and human adenovirus. The primer probe group for detecting G II type norovirus provided by the invention is high in sensitivity, and the lowest detection limit is 1.22 copies/microliter.

Description

technical field [0001] The invention belongs to the technical field of norovirus detection, and in particular relates to a primer probe set, a kit and a method for detecting GII type norovirus. Background technique [0002] Norovirus is the main cause of non-bacterial gastroenteritis, which leads to large-scale outbreaks. Norovirus is a single-stranded positive-sense RNA virus with a diameter of about 27 nm and icosahedral symmetry. The full length of the norovirus genome is about 7.6kb, encoding three open reading frames (Open reading frame, ORF). Open reading frame 1 (ORF1) encodes non-structural proteins, which are cleaved into six proteins after translation, including RNA-dependent RNA polymerase (RNA-dependent RNA polymerase, RdRp). Open reading frame 2 (ORF2) and open reading frame 3 (ORF3) encode the major capsid protein (Viral protein 1, VP1) and the minor capsid protein (Viral protein 2, VP2), respectively. Noroviruses are genetically and antigenically diverse. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6844C12N15/11C12R1/93
CPCC12Q1/701C12Q1/6844C12Q2521/507C12Q2522/101C12Q2531/119
Inventor 周树青李君文杨栋谌志强金敏尹静陈郑珊师丹阳李海北杨忠委陈天姣王华然
Owner INST OF ENVIRONMENTAL MEDICINE & OCCUPATIONAL MEDICINE ACAD OF MILITARY MEDICINE ACAD OF MILITARY SCI
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