Primer group for detecting influenza a H1N1 virus in respiratory secretions of children

A technology of H1N1 and influenza virus, which is applied in the detection/inspection of microorganisms, microorganisms, recombinant DNA technology, etc., can solve the problems of expensive equipment, low sensitivity and detection rate, high technical requirements, etc., and achieve high sensitivity, Easy to promote and easy to operate

Inactive Publication Date: 2019-08-09
SHANGHAI CITY JIADING DISTRICT CENT HOSPITAL
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  • Description
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Problems solved by technology

Virus isolation takes days to weeks to obtain results, which is not suitable for its clinical detection during influenza epidemics, and requires a highly complex biosafety laboratory; although the cycle of serological detection (antigen / antibody detection) can meet clinical requirements The need for timeliness, but the sensitivity and detection rate are too low, so it can only be used as an auxiliary method
[0006] Although conventional PCR and real-time quantitative PCR techniques are currently widely used molecular biology detection methods, they are not suitable for on-site rapid detection and Grassroots popularization and application

Method used

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  • Primer group for detecting influenza a H1N1 virus in respiratory secretions of children
  • Primer group for detecting influenza a H1N1 virus in respiratory secretions of children
  • Primer group for detecting influenza a H1N1 virus in respiratory secretions of children

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Embodiment Construction

[0023] The present invention is further described below.

[0024] Operation steps of the present invention are:

[0025] 1. Extract RNA;

[0026] Viral RNA was extracted from the secretion using the GeneJET Viral DNA / RNA Purification Kit (Thermo Fisher Scientific, USA) according to the instructions.

[0027] 2. Primer sequence:

[0028]

[0029] 3. Use the RNA extracted in step 1 as a template to perform RT-LAMP amplification reaction, and the total reaction volume of the RT-LAMP detection reaction system is 25 μl.

[0030] 4. The RT-LAMP detection reaction system is:

[0031] dNTPs(1.4mM),Tris-HCl(PH8.8,20mM),(NH4)2SO4(10mM),KCl(10mM),MgSO 4 (8mM), betaine (0.8M), 0.1% Tween20, Bst DNA polymerase (8 U), AMV reverse transcriptase (0.5U), fluorescent dye (SYBR green I1μl), H1N1 viral RNA (2μl). Amplify at 63 degrees Celsius for 1 hour, and place the reaction tube in a water bath at 80 degrees Celsius for 5 minutes to terminate the reaction. Observe the green fluorescen...

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Abstract

The invention provides a primer group for detecting the influenza a H1N1 virus in respiratory secretions of children, and further relates to a fast quantitative detection method using the primer groupand a fast quantitative detection kit containing the primer group. The primer group is composed of an outer primer pair F3 / B3 and an inner primer pair FIP / BIP. The detection method and the kit are easy to operate, fast, accurate, easy to popularize and high in sensitivity, no expensive instruments are needed, the detection method and the detection kit can be used in detection of on-site pathogensand primary hospitals, and the primer group has an important significance for monitoring and controlling the influenza a virus of the children.

Description

technical field [0001] The invention belongs to the technical field of virus detection, and in particular relates to a primer set for rapid and quantitative detection of influenza A (H1N1) virus RNA in children's respiratory secretions. [0002] The present invention also relates to a method for detecting influenza A (H1N1) virus in children's respiratory secretions by using the above-mentioned primer set. [0003] The present invention also relates to a rapid quantitative detection kit for detecting influenza A H1N1 virus in children's respiratory secretions by using the above primer set. Background technique [0004] The clinical symptoms of influenza and other respiratory infections can be so similar that it is difficult to distinguish them, and only a laboratory can provide an accurate diagnosis of influenza viruses. Influenza A (H1N1) is the most common outbreak that is easily affected by seasons. Timely and accurate identification of virus species is of great signific...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6851C12N15/11C12R1/93
CPCC12Q1/701C12Q1/6851C12Q2531/119C12Q2521/107
Inventor 沈丽娟常庆孙杰王飞史俊峰徐炜新于修文沙爽
Owner SHANGHAI CITY JIADING DISTRICT CENT HOSPITAL
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