Method for detecting nucleic acids by using dual real-time fluorescent isothermal amplification technology

A real-time fluorescence and isothermal amplification technology, which is applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganism measurement/testing, etc., can solve the problems of inapplicable clinical diagnosis, poor specificity and sensitivity, false positives, etc.

Active Publication Date: 2020-07-10
鲲石生物科技(深圳)有限公司
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Problems solved by technology

CT scanning technology is convenient but not specific enough, which can assist in the diagnosis; immunochromatography is simple to operate, but has poor specificity and sensitivity, which can easily cause false negative and false positive results, and is not suitable for clinical diagnosis; High sensitivity, strong specificity, suitable for clinical diagnosis and diagnosis of new coronavirus

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  • Method for detecting nucleic acids by using dual real-time fluorescent isothermal amplification technology
  • Method for detecting nucleic acids by using dual real-time fluorescent isothermal amplification technology
  • Method for detecting nucleic acids by using dual real-time fluorescent isothermal amplification technology

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[0034] A method for detecting nucleic acids using dual real-time fluorescence isothermal amplification technology. The detection method for synchronous detection of new coronavirus SARS-CoV-2 and influenza A H1N1 virus using dual real-time fluorescence isothermal platform technology includes the following steps:

[0035] (1) Instruments and reagents

[0036] Fluorescence quantitative PCR instrument (Bio-Rad CFX96 touch),

[0037] PCR tubes that are nuclease-free and have light-transmitting PCR caps,

[0038] 0-2 μL pipette (Eppendorf),

[0039] 1-10 μL pipette (Eppendorf),

[0040] 2-20 μL pipette (Eppendorf),

[0041] 10-100 μL pipette (Eppendorf),

[0042] 20-200 μL pipette (Eppendorf),

[0043] 200-1000 μL pipette (Eppendorf),

[0044] Viral RNA reverse transcription kit (SuperScriptTM III First-Strand Synthesis System, ThermoFisher),

[0045] DNA Extraction Kit (E.Z.N.A.® Gel Extraction Kit, OMEGA BIO-TEK),

[0046] TA Cloning Kit (pEASY®-T1 Cloning Kit, TransGen ...

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Abstract

The invention belongs to the technical field of virus detection and discloses a method for detecting nucleic acids by using a dual real-time fluorescent isothermal amplification technology. By adopting the method, existence of RNA of a novel corona virus SARS-VoV-2 and an influenza A H1N1 virus is simultaneously detected through a double-channel double-florescent channel isothermal platform. The method has the advantage of multiplicity, is capable of detecting two viruses in one reaction system, is short in detection cycle, is applicable to rapid clinical detection and diagnosis, and is high in virus detection specificity, high in accuracy, high in detection sensitivity, good in experiment result repeatability and high in precision; and a detection primer has specificity, the detection efficiency can be greatly improved, the amount of reagents in amplification systems can be reduced, and the detection cost can be reduced.

Description

technical field [0001] The invention belongs to the technical field of virus detection and discloses a method for detecting nucleic acid by using double real-time fluorescence isothermal amplification technology. Background technique [0002] Corona Virus Disease 2019 (COVID-19), referred to as "New Coronary Pneumonia", named by the World Health Organization as "2019 Coronavirus Disease", refers to the pneumonia caused by the 2019 Novel Coronavirus (SARS-Cov-2) infection , which was identified as a beta-type coronavirus (betacoronavirus) containing single-stranded positive-strand RNA (ssRNA), its structure is very similar to severe acute respiratory syndrome SARS, and the new coronavirus gene has similarities with SARSr-CoV and MERSr-CoV The sequence similarity is about 70% and 40%, and the whole genome sequence identity with the bat virus TG13 is as high as 96%. The main sequence of nucleic acid includes E (envelope protein gene), M (membrane protein gene), N (nucleocapsid...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6844C12Q1/70C12R1/93
CPCC12Q1/6844C12Q1/701C12Q2600/16C12Q2537/143C12Q2563/107C12Q2531/119C12Q2521/107Y02A50/30
Inventor 尹秀山于琳
Owner 鲲石生物科技(深圳)有限公司
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