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Kit for detecting GII type norovirus and applications thereof

A kit and virus technology, applied in the field of molecular biology and virus detection, can solve the problems of low accuracy and sensitivity, difficulty in improving detection speed, precision and expensive instruments, etc., achieve low reaction temperature, constant reaction temperature, and short detection time Effect

Inactive Publication Date: 2013-11-20
SHANGHAI OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The strain-type specificity of the ELISA detection method is too strong, the accuracy and sensitivity are low, and the detection is unstable, so the application range is relatively narrow, and it is difficult to meet the requirements of modern detection and diagnosis technology.
Although the PCR method (including RT-PCR and Real-time RT-PCR) makes up for the shortcomings of ELISA, the main technical factor that limits its application is that the instrument is sophisticated and expensive, and depends on precise variable temperature conditions, so this method can only be limited to professional The use of the laboratory, and the detection speed is difficult to improve
The LAMP method needs to react at 65°C for 45-90 minutes, which is simple and lacks portability
However, there are no relevant reports on primers and probes combined with RPA technology for the detection of GII type norovirus.

Method used

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  • Kit for detecting GII type norovirus and applications thereof
  • Kit for detecting GII type norovirus and applications thereof

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Experimental program
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Effect test

Embodiment 1

[0019] Embodiment 1 Design and verification of primers and probes of the present invention

[0020] 1. Experimental materials

[0021] Reverse transcriptase and 1×RT reaction buffer, produced by Roche; RNaseOut inhibitor, produced by Invitrogen; TwistAmp DNA amplification kit produced by TwistDx, including Rehydration Buffer reaction buffer solution, DNA recombinase, DNA polymerase, single Chain binding protein, phosphokinase, ATP and other reagents, among which DNA recombinase, DNA polymerase, single chain binding protein, phosphokinase, ATP and other reagents are stored in the reaction centrifuge tube in the form of dry powder.

[0022] 2. Experimental method

[0023] 2.1 Design of primers and probes

[0024] According to the target sequence of Norovirus type GII (NCBI accession number: X86557), the following primers and probes were designed to specifically recognize the target sequence:

[0025] Forward primer (FP2): 5'-TGGATGAGATTCTCAGATCTGAGCACGTGGGAGGG-3' (SEQ ID NO...

Embodiment 2

[0046] Embodiment 2 Kit 1 of the present invention

[0047] The kit contains the following reagents and materials:

[0048] Primer solution: forward primer FP2 (SEQ ID NO.1, 5 μmol / L); reverse primer RP3 (SEQ ID NO.2, 5 μmol / L).

Embodiment 3

[0049] Embodiment 3 Kit 2 of the present invention

[0050] The kit contains the following reagents and materials:

[0051] Primer solution: forward primer FP2 (SEQ ID NO.1, 5 μmol / L); reverse primer RP3 (SEQ ID NO.2, 5 μmol / L); probe (SEQ ID NO.3, 5 μmol / L).

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Abstract

The invention relates to a kit for detecting GII type norovirus and applications thereof, concretely a primer pair and a probe for detecting the GII type norovirus, and a kit comprising the primer pair and the probe. The GII type norovirus in a sample cab be successfully detected by a recombinase polymerase amplification reaction, and can be quantified by fluorescence intensity. The kit has advantages of high sensitivity, strong singularity, short detection time, and constant and low reaction temperature, can be used for qualitative screening and quantitative determination for GII type norovirus in the sample, such as a clinic sample, foodstuff, an aquatic product, a water body, etc. accordingly, the kit can be prepared to be a diagnostic kit for GII type norovirus.

Description

technical field [0001] The invention relates to the technical fields of molecular biology and virus detection, in particular to a kit for detecting GII type norovirus and its application. Background technique [0002] Norovirus is the most important pathogen causing non-bacterial gastroenteritis outbreaks in the world. It is highly infectious and can cause small-scale outbreaks or large-scale epidemics with up to hundreds of thousands of infections. It can infect all ages Group of people. Norovirus is a virus with large genetic variation. It can be divided into five groups (Genogroup G) according to its antigenicity and nucleotide sequence variation. GI, GII, and GIV can infect humans, of which GI and GII are infectious The main genetic groups of humans can be further subdivided into different genotypes (GI.1, GI.2, GI.3, GI.4, GI.5, GI.6, GI.7, GI.8, GII.1, GII.2, GII.3, GII.4, GII.5, GII.6, GII.7, GII.8, GII.9, GII.10, GII.11, GII. 12. GII.13, GII.14, GII.15, GII.16, GI...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68C12N15/11
Inventor 王永杰喻勇新严淑玲潘迎捷
Owner SHANGHAI OCEAN UNIV
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