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Novel coronavirus detection kit

A detection kit, a technology for coronavirus, which is used in the determination/inspection of microorganisms, biochemical equipment and methods, microorganisms, etc., can solve the problems of irregular sample collection, high false negative test results, and short research and development cycle, and achieve the preparation process. Simple and safe, high detection accuracy, and the effect of improving the detection rate

Pending Publication Date: 2020-05-22
SICHUAN MACCURA BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Due to the short development cycle and irregular sample collection, the current novel coronavirus detection kits generally have the problem of high false negative detection and easy missed detection; and the safety of detection kits is another problem that needs to be solved urgently

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] A coronavirus detection kit, said kit comprising qRT-PCR reaction solution, qRT-PCR enzyme solution, internal standard, negative quality control and positive quality control;

[0055] The qRT-PCR reaction solution: primers, probes, dNTP, Mg2+, PBS buffer;

[0056] The qRT-PCR enzyme liquid: Taq enzyme, UDG enzyme;

[0057] The internal standard: RNA pseudovirus containing internal reference detection gene fragments;

[0058] The negative quality control product: DEPC-treated water;

[0059] The positive quality control product: RNA pseudovirus containing the target detection gene fragment;

[0060] The primers and probes of this kit are as follows:

[0061] The primer sequence for detecting the ORF1ab gene of the 2019-nCoV virus is SEQ ID NO: 1-2, and the probe sequence is SEQ ID NO: 9;

[0062] The primer sequence for detecting the N gene of the 2019-nCoV virus is SEQ ID NO: 3-4, and the probe sequence is SEQ ID NO: 10;

[0063] The primer sequence for detecting t...

Embodiment 2

[0067] Detection method of coronavirus detection kit

[0068] Reagent preparation

[0069] Take each component of the nucleic acid extraction or purification reagent in the kit, place it at room temperature, and wait for the temperature to reach room temperature, mix well and centrifuge immediately for later use.

[0070] Prepare qRT-PCR mix according to the following table:

[0071] Table 2

[0072] qRT-PCR reaction solution qRT-PCR enzyme solution Number of tests qRT-PCR Mixture 18μL 2μL N=n+2

[0073] Wherein, the number of tests N=n+2, wherein, n represents the number of samples to be tested, and the other 2 are negative quality control products and positive quality control products. According to the laboratory situation, due consideration should be given to the calculation during the process of dispensing the mixed solution.

[0074] Transfer the above prepared reagents to the sample processing area

[0075] sample processing

[0076] ...

Embodiment 3

[0091] The internal standard and positive quality control were verified using the detection method of Example 2.

[0092] The IC09 pseudovirus particles (internal standard) of Sichuan New Materials were diluted 10 times with the pseudovirus diluent, and the 2019-nCoV nucleic acid detection reagent of Mike Bio was used for multiple amplification verification. The Ct values ​​of each channel are shown in the following table:

[0093] Table 4

[0094]

[0095] The CoV-2019 pseudovirus particles (positive quality control) of Sichuan New Materials were diluted 10 times with the pseudovirus diluent, and the nucleic acid extraction or purification kit and 2019-nCoV nucleic acid detection reagent of Mike Bio were used for amplification verification. The test result was positive, and the results are as follows:

[0096] table 5

[0097]

[0098] In the sample, 3L~9L means that 10 3 diluted, 10 4 diluted, 10 5 Dilution 10 6 diluted, 10 7 diluted, 10 8 diluted, 10 9 diluti...

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Abstract

The invention discloses a novel coronavirus detection kit. The kit comprises a qRT-PCR reaction liquid, a qRT-PCR enzyme liquid, an internal standard, a negative quality control substance and a positive quality control substance, wherein the internal standard is an RNA pseudovirus containing an internal reference detection gene fragment; the positive quality control substance is an RNA pseudoviruscontaining a target detection gene fragment; and the qRT-PCR reaction liquid comprises four groups of upstream and downstream primers with sequences of SEQ ID NO:1-8 and four probes with sequences ofSEQ ID NO:9-12. The kit can be used to simultaneously detect three sections of target gene fragments, so that missed detection can be effectively avoided. In addition, pseudoviruses are used in boththe internal standard and the positive quality control substance of the kit, so that safety of the kit is effectively improved, and detection accuracy of the kit is further ensured. The kit is high sensitivity and good in stability.

Description

technical field [0001] The invention belongs to the field of in vitro nucleic acid detection. The invention provides a fluorescent RT-PCR detection kit for detecting novel coronavirus (2019-nCoV) in clinical samples; it can be used for early clinical qualitative diagnosis of novel coronavirus. Background technique [0002] Coronavirus is a large virus family that exists widely in nature. It is named for its shape that looks like a crown when observed under an electron microscope. It mainly causes respiratory diseases. On February 11, 2020, the World Health Organization (WHO) announced that it will officially name the new type of coronavirus pneumonia that began to appear in December 2019 as "COVID-19", the full name of which is CoronavirusDisease 2019. After being infected with the new coronavirus, the initial symptoms of patients are mostly fever, fatigue, and dry cough, and they gradually develop severe symptoms. Some severe cases may develop acute respiratory distress syn...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/686C12R1/93
CPCC12Q1/701C12Q1/686C12Q2600/166C12Q2563/107C12Q2521/107
Inventor 杨正林杨文秀石毅蒋黎龙腾镶杨达梅文春描占明明
Owner SICHUAN MACCURA BIOTECH CO LTD
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