Reagent kit for detecting novel coronaviruses and influenza viruses

A technology for coronaviruses and influenza viruses, applied in the field of molecular biology detection, to achieve good specificity, avoid false positives or false negatives, and high sensitivity

Active Publication Date: 2020-06-09
3D BIOMEDICINE SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, PCR detection kits for influenza virus detection and PCR detection kits for novel coronavirus COVID-19 are already on the market, but the sensitivity and accuracy of COVID-19 detecti

Method used

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  • Reagent kit for detecting novel coronaviruses and influenza viruses
  • Reagent kit for detecting novel coronaviruses and influenza viruses
  • Reagent kit for detecting novel coronaviruses and influenza viruses

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Embodiment 1 Primer probe, reagent and detection method

[0063] 1.1 Primer probe sequence

[0064] The ORF1ab gene of the novel coronavirus COVID-19 was selected as the amplification target region, specific primers and fluorescent probes were designed, and the probes were labeled with FAM to detect whether the samples contained novel coronavirus RNA. Select the S gene of the new coronavirus, design specific primers and fluorescent probes, and the probes are labeled with ROX to detect whether the sample contains the new coronavirus RNA. The characteristic conserved sequence of influenza virus A was selected, specific primers and fluorescent probes were designed, and the probe was labeled with Cy5 to detect whether the sample contained influenza virus ARNA. The characteristic conserved sequence of influenza virus B was selected, specific primers and fluorescent probes were designed, and the probe was labeled with VIC to detect whether the sample contained influenza viru...

Embodiment 2

[0095] Embodiment 2 detection sensitivity

[0096] The sensitivity was verified using the primer-probe combination (Table 2), reagents, and nucleic acid amplification detection method in Example 1.

[0097] For COVID-19, the RNA extracted from 3 clinical real samples was mixed into 1 part. After quantification by ddPCR, it was 6460copeis / μL, and the samples were diluted to 8 copies / μL, 0.8copies / μL, and 0.4copies / μL with virus preservation solution. μL, 0.2copies / μL, 0.1 copies / μL, 0.05 copies / μL, for nucleic acid extraction and amplification.

[0098] S4 (S4:inFluA / H3N2, 1.2×10 4 copies / μL) were evaluated, and the virus preservation solution was used to perform 10-fold serial dilutions, and were diluted to 1.2×10 3 Copies / μL, 120copies / μL, 12copies / μL, 1.2copies / μL, 0.12copies / μL, amplified after extraction.

[0099] Influenza virus B (influB) uses S1 of the national reference product of the second-generation influenza A / B nucleic acid detection reagent (S1:inFluB / Victoria...

Embodiment 3

[0101] Example 3 specificity

[0102] The specificity was verified by using the primer-probe combination (Table 2), reagents and nucleic acid amplification detection method in Example 1.

[0103] In order to prove the cross-reactivity of the kit, human coronavirus OC43, human coronavirus 229E, human coronavirus NL63, human coronavirus HKU1, SARS coronavirus, MERS coronavirus, meningococcus, Haemophilus influenzae, gold Staphylococcus aureus, Streptococcus pneumoniae, Rubella virus, Mumps virus, Respiratory adenovirus type 3, Respiratory adenovirus type 7, Respiratory syncytial virus type B, Parainfluenza virus type 2, Streptococcus pyogenes, Bordetella pertussis, Candida albicans, Pseudomonas aeruginosa, Staphylococcus epidermidis and other 21 pathogens and human genome DNA were detected respectively.

[0104] Firstly, the virus was diluted to 1E+05 CFU / mL, and the bacterial and human genomic DNA was diluted to 1E+06 copies / mL, and then the nucleic acid of each pathogenic mic...

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Abstract

The invention discloses a reagent kit for detecting novel coronaviruses and influenza viruses at the same time. The reagent kit disclosed by the invention specially comprises a primer probe for separately targeting S genes, E genes, N genes and/or ORF1ab genes of novel coronaviruses, and a primer probe for targetting characteristic consensus sequence of the influenza viruses A/B, and novel coronavirus RNA for qualitatively detecting pneumonia suspected cases and suspected clustered cases of patients infected with the novel coronaviruses, or other novel coronavirus RNA for diagnosing infectionof the novel coronaviruses or identifying novel coronavirus RNA in other samples of nasopharyngeal swabs, sputamentum and the like of diagnostician, and besides, the reagent kit is used for detectingRNA of influenza viruses A/B in the samples to assist in elimination of suspected cases, can be used for identifying and diagnosing infection with the novel coronaviruses, and is high in sensitivity and convenient to use.

Description

technical field [0001] The invention relates to molecular biology detection technology, in particular to a detection kit for detecting novel coronavirus and / or influenza virus and its application method. Background technique [0002] The 2019 novel coronavirus (COVID-19) belongs to the genus Betacoronavirus and is the seventh human-infecting coronavirus (HCoV) identified in human history. Its genetic characteristics are significantly different from those of SARS-CoV and MERS-CoV . On February 11, 2020, the International Committee on Taxonomy of Viruses (ICTV) announced that the classification of the 2019 novel coronavirus was named Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-Cov-2). The disease caused by viral infection is officially named COVID-19. The 2019 novel coronavirus pneumonia is mainly transmitted through droplets and contact, and the crowd is generally susceptible. The general symptoms of 2019-nCoV infection are: fever, fatigue, dry cough, and gradual ...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6851C12N15/11C12R1/93
CPCC12Q1/6851C12Q1/701C12Q2600/166C12Q2531/113C12Q2521/107C12Q2545/101C12Q2563/107
Inventor 周科隆仁青措张华曹渊王晓楠熊磊陈才夫
Owner 3D BIOMEDICINE SCI & TECH CO LTD
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