Dual fluorescence quantification RT-PCR detection kit and application thereof

A technology of RT-PCR and detection kits, which is applied in the direction of fluorescence/phosphorescence, microbial measurement/inspection, biochemical equipment and methods, etc., can solve the problems of wasting clinical specimens and reagents, time-consuming and laborious, and save clinical specimens and Effects of Reagents, Improving Influenza Testing, Increased Biosecurity Risk

Active Publication Date: 2011-11-02
ZHEJIANG CENT FOR DISEASE CONTROL & PREVENTION
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  • Abstract
  • Description
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  • Application Information

AI Technical Summary

Problems solved by technology

[0008] In view of the increasing importance of human seasonal influenza, avian influenza and H1N1 influenza in public health, it is time-consuming, laborious and wasteful of clinical specimens and reagents to detect multiple influenza viruses at the same time during influenza surveillance

Method used

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  • Dual fluorescence quantification RT-PCR detection kit and application thereof
  • Dual fluorescence quantification RT-PCR detection kit and application thereof

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Embodiment 1

[0041] 1. Materials and methods

[0042] Virus strains and clinical specimens:

[0043] Influenza A H1N1, SWH1N1, seasonal H1N1, and seasonal H3N1 virus strains were isolated from Zhejiang Provincial Center for Disease Control and Prevention. The clinical samples were obtained from the throat swabs of suspected patients with the recent outbreak of H1N1 virus in Zhejiang Province. The samples were collected and transported to the laboratory with ice.

[0044] 1.2 Primers and probes

[0045] A(H1N1), seasonal H1N1, porcine H1N1 influenza virus strains from all over the world were downloaded from the NCBI gene bank in the United States. Homology comparisons were carried out, and influenza A H1N1-specific primers and Taqman probes were designed in the HA gene region of all influenza virus genomes. The sequences are as follows:

[0046] Upstream primer alpha-infA-FP: 5’- GACCRATCYTGTCACCTCTGAC-3’

[0047] Downstream primer alpha infA-RP: 5'-AGGGCATTYTGGACAAAKCGTCTA-3'

[0048]...

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Abstract

The invention provides a dual fluorescence quantification RT-PCR (reverse transcription-polymerase chain reaction) detection kit, comprising a deoxynucleoside triphosphate mixture, MgCl2, an RNA enzyme inhibitor, a Moloney murine leukemia virus reverse transcriptase, a DNA polymerase, a influenza virus standard and a reference substance. Based on sequence analysis of present pervasive A H1N1 influenza virus, the invention provides a multiple fluorescence quantification PCR molecular biology gene diagnosis method and a diagnostic kit which are rapid, specific, accurate and sensitive. In addition, one reaction tube can simultaneously detect and differentiate an influenza A virus or an influenza B virus in 2h, so as to improve influenza detection.

Description

technical field [0001] The invention belongs to the field of biological technology, and relates to a double fluorescence quantitative RT-PCR detection kit for influenza A virus and influenza B virus nucleic acid and an application thereof. Background technique [0002] Since March 18, 2009, there has been an outbreak of human infection with H1N1 influenza virus in Mexico. Since the first death case of human infection with influenza A H1N1 virus appeared in Mexico on April 13, 2009, as of May 5, 21 countries around the world have confirmed the existence of influenza A (H1N1) influenza cases through laboratory testing. The number of cases rose to 1,124, including 26 deaths. In addition to Mexico and the United States, many countries have reported confirmed cases of influenza A (H1N1). To this end, the World Health Organization (WHO) raised the global influenza pandemic warning level to level 4, and further raised it to level 5 on April 29, which means that in one of the WHO'...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68G01N21/64
Inventor 张严峻
Owner ZHEJIANG CENT FOR DISEASE CONTROL & PREVENTION
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