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Nucleic acid detection reagent kit for novel coronavirus 2019-nCoV

A 2019-ncov, coronavirus technology, applied in the field of biotechnology and molecular diagnosis, can solve the problems of lack of simultaneous detection of multiple genes, unfavorable virus detection and diagnosis, and large differences in reagent performance

Pending Publication Date: 2020-06-05
DAAN GENE CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] There are currently some nucleic acid detection kits developed based on real-time fluorescent quantitative PCR technology on the market, but most of them are single-target detection reagents, lacking simultaneous multi-gene detection to confirm the virus, and the performance of the reagents varies greatly, which is not conducive to the detection of the virus and diagnosis

Method used

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  • Nucleic acid detection reagent kit for novel coronavirus 2019-nCoV
  • Nucleic acid detection reagent kit for novel coronavirus 2019-nCoV
  • Nucleic acid detection reagent kit for novel coronavirus 2019-nCoV

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0118] Example 1 Detection limit test of novel coronavirus 2019-nCoV nucleic acid detection kit

[0119] Dilute the pseudovirus with the determined value as the initial sample to a concentration of 10 5 copies / ml, serially diluted to 10 4 、10 3 , 500, 250copies / ml, each concentration sample was added to the final concentration of 10 4 Copies / ml of the pseudovirus containing the amplified fragment of the internal standard is used as the sample to be tested, and the sensitivity of the 3-fold detection reagent is tested. The PCR reaction system comprises the combination of primers and probes shown in SEQ ID NO.: 1-12 of the present invention.

[0120] For test results, see Figure 1-4 :

[0121] figure 1 : N gene detection limit;

[0122] figure 2 : ORF1ab gene detection limit test;

[0123] image 3 : E gene detection limit test;

[0124] Figure 4 : internal standard test result.

[0125] The results showed that the lowest detectable concentration was 500copies / ...

Embodiment 2

[0126] Example 2 Specificity Test of New Coronavirus 2019-nCoV Nucleic Acid Detection Kit

[0127] Other pathogens that are similar to the 2019 new coronavirus species or cause similar symptoms (such as seasonal influenza A H1N1 virus, new type A H1N1 (2009) influenza virus, influenza A H3N2, H5N1, H7N9, influenza B Yamagata, Influenza B Victoria, RSV A, RSV B, parainfluenza virus, adenovirus, enterovirus, human metapneumovirus (human metapneumovirus), Epstein-Barr virus, measles virus, human giant Cytovirus, rotavirus, norovirus, mumps virus, varicella-zoster virus, Mycoplasma pneumoniae, Chlamydia pneumoniae, Legionella, Bordetella pertussis, Haemophilus influenzae, Staphylococcus aureus, Streptococcus pneumoniae, pyogenic Streptococcus, Klebsiella pneumoniae, Mycobacterium tuberculosis, Aspergillus fumigatus, Candida albicans, Candida glabrata, Cryptococcus neoformans, coronavirus (HKU1, OC43, NL63, 229E) and human genomic DNA as specific reference materials, Test the spec...

Embodiment 3

[0132] Example 3, Precision Test of Novel Coronavirus 2019-nCoV Nucleic Acid Detection Kit

[0133] N / ORF1ab / E gene pseudovirus was diluted to 10 5 and 10 3 Copies / ml was used as a precision reference product, each repeated 10 times, and the coefficient of variation of each concentration precision reference product was calculated.

[0134] For test results, see Figure 7-9 :

[0135] Figure 7 . N gene precision test results;

[0136] Figure 8 . ORF1ab gene precision test results;

[0137] Figure 9 .E gene precision test results;

[0138] After testing, the coefficients of variation of the new coronavirus 2019-nCoV high-concentration and low-concentration precision reference products are 1.16% and 1.54% for the N gene, 1.16% and 1.54% for the ORF1ab gene, and 1.16% and 1.54% for the E gene; The coefficients of variation of precision reference products with different concentrations of detection targets were all less than 5%.

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Abstract

The invention provides a bucleic acid detection reagent kit for novel coronavirus 2019-nCoV, and particularly relates to a reagent kit and method for multiple detection of nucleic acid of novel coronavirus 2019-nCoV. 3 nucleic acid targets of the novel coronavirus 2019 nCoV can be detected at the same time, through mutual authentication between different targets, false positive can be prevented, and undetection which can be possibly caused by mutation is confirmed, so that the virus identification accuracy is notably improved.

Description

technical field [0001] The invention belongs to the fields of biotechnology and molecular diagnosis, and in particular, the invention relates to a primer-probe combination and a kit for detecting the novel coronavirus 2019-nCoV. Background technique [0002] Coronaviruses belong to the genus Coronavirus in the family Coronaviridae. The virus of the genus Coronavirus is a positive-stranded single-stranded RNA virus with a mantle, with a diameter of about 80-120 nm. Its genetic material is the largest among all RNA viruses, infecting humans, mice, pigs, cats, dogs, wolves, chickens, and cattle. , avian vertebrates. A variant of coronavirus is the pathogen that causes SARS and is an RNA virus. The diameter of the virus particle is 60-200nm, the average diameter is 100nm, it is spherical or elliptical, and it is polymorphic. The virus has an envelope, and there are spines on the envelope. The whole virus is like the corona, and the spines of different coronaviruses are obviou...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/686C12N15/11
CPCC12Q1/701C12Q1/686C12Q2600/16C12Q2600/166C12Q2537/143C12Q2545/113C12Q2521/107C12Q2545/101Y02A50/30C12Q1/6883C12Q1/6876
Inventor 蒋析文范建彭海龙
Owner DAAN GENE CO LTD
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