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Hantavirus ultra-fast fluorescent PCR detection kit and primer probe combination thereof

A detection kit and technology for Hantavirus, applied in the field of biotechnology applications, can solve the problems of long detection period, low detection sensitivity of Hantavirus, low detection throughput, etc., achieve good accuracy, avoid non-specific amplification, The effect of high detection sensitivity

Inactive Publication Date: 2019-02-19
南通国际旅行卫生保健门诊部
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  • Abstract
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Problems solved by technology

[0005] The purpose of the present invention is to provide a hantavirus extremely fast fluorescent PCR detection kit and its primer probe combination to solve the problems of low detection sensitivity, long detection cycle and low detection throughput of hantavirus in the prior art

Method used

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  • Hantavirus ultra-fast fluorescent PCR detection kit and primer probe combination thereof
  • Hantavirus ultra-fast fluorescent PCR detection kit and primer probe combination thereof
  • Hantavirus ultra-fast fluorescent PCR detection kit and primer probe combination thereof

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Embodiment 2

[0060] Example 2 Hantavirus Detection Operation Specification 1. Nucleic Acid Extraction

[0061] 1.1 Nucleic acid extraction: The kit is equipped with DNA / RNA one-step lysis and extraction reagents. Add 200 μL serum sample to 50ul lysate, centrifuge at 5000 RPM for 2 minutes at 95°C for 2 minutes, and take 50 μL of the supernatant as a template. For specific extraction steps, please refer to the corresponding extraction kit manual. .

[0062] 2. Reagent preparation

[0063] 2.1 After dissolving the amplification reagent at room temperature, according to the number of samples to be tested (n), prepare the amplification reagent according to the number (n+2), divide 8 μL per person, negative and positive control reagents, briefly centrifuge after melting at room temperature;

[0064] 2.2 Move the above-mentioned 16-well chip plate to the sample processing area.

[0065] 3. Adding samples

[0066] Use a suction nozzle with a filter element to take 2 μL of the supernatant of th...

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Abstract

The invention relates to a Hantavirus ultra-fast fluorescent PCR detection kit and a primer probe combination thereof. The Hantavirus ultra-fast fluorescent PCR detection kit comprises an extraction reagent, an amplification reagent and a control reagent; the extraction reagent is composed of sodium dodecyl sarcosinate|sodium N-Lauroyl sarcosinate (NLS) [(W / V)NLS], dithiothreitol DTT, TE buffer liquid and an NP-40 surfactant; the amplification reagent includes an RT-PCR reaction solution mixed with Hantavirus and a detection reagent of a 16-pore microfluidic chip; the control reagent comprisesa negative control and a positive control; the detection reagent comprises the RT-PCR reaction solution and is composed of Buffer, 2.0 mM dNTPs, 1 U / MuL of Taq DNA polymerase, 2 U / MuL of M-MLV reverse transcriptase, 0.3 U / MuL of RRI, the Hantavirus and a human internal standard gene detection primer probe. The negative control includes DEPC water, and the positive control includes artificially constructed pseudo-viruses containing fragments of target genes.

Description

technical field [0001] The invention discloses a hantavirus extremely fast fluorescent PCR detection kit and a primer-probe combination thereof, belonging to the field of biotechnology applications. Background technique [0002] Hantavirus belongs to the Bunyaviridae family and is a negative-strand RNA virus with enveloped segments. The genome includes L, M, and S3 segments, which encode L polymerase protein, G1 and G2 glycoproteins, and nucleoproteins, respectively. . [0003] There are two types of hantavirus: one causes hantavirus pulmonary syndrome (HPS) and the other causes hantavirus hemorrhagic fever with renal syndrome (HFRS). The former is mainly endemic in the United States, but cases have also been found in Argentina, Brazil, Paraguay, Bolivia, and Germany. The main clinical manifestations are acute respiratory failure characterized by non-cardiogenic pulmonary edema and high fatality rate (52.4% to 78.0%) after prodromal symptoms such as fever and headache for ...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/686C12Q1/6806C12N15/11
CPCC12Q1/6806C12Q1/686C12Q1/701C12Q2600/166C12Q2521/107C12Q2563/107C12Q2545/101C12Q2561/101C12Q2527/125
Inventor 陈峰杨庆贵杨国平唐晓宇
Owner 南通国际旅行卫生保健门诊部
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