Novel coronavirus nucleic acid detection kit

A technology for detection kits and coronaviruses, applied in the determination/inspection of microorganisms, biochemical equipment and methods, microorganisms, etc., can solve problems such as complex operations, unsuitable diagnostic detection methods, and long time consumption, and achieve high sensitivity and difficulty Contamination, high specific effect

Pending Publication Date: 2021-08-27
HANGZHOU MATRIDX BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, this method is complicated to operate, takes a long time, requires a sophisticated sequencer, and is expensive to sequence, so it is not suitable as a routine diagnostic method at present.

Method used

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  • Novel coronavirus nucleic acid detection kit
  • Novel coronavirus nucleic acid detection kit
  • Novel coronavirus nucleic acid detection kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] Embodiment 1: the detection of kit sensitivity of the present invention

[0082] 1. Preparation for detection of sample nucleic acid:

[0083] 1) Take the concentration of 30,000 copies / mL after the inactivation of the culture of the new coronavirus as the S1 sample, and sequentially dilute it with PBS according to a ratio of 1:3, and then the S2 sample is 10,000 copies / mL, the S3 sample is about 3,000 copies / mL, and the S4 sample is about 3000 copies / mL. 1000copies / mL, S5 sample is about 300copies / mL, S6 sample is about 140copies / mL, S7 sample is about 40copies / mL, S8 sample is about 15copies / mL, NC sample is PBS solution. After nucleic acid extraction, the above S1-S8 samples and NC samples become the nucleic acid sample solution to be detected by the kit in this embodiment. The process of nucleosomes uses the pathogenic biological nucleic acid extraction kit (article number MD002) provided by our company (Hangzhou Jieyi Biotechnology Co., Ltd.).

[0084] 2) Through...

Embodiment 2

[0111] Embodiment 2: the detection of kit specificity of the present invention

[0112] 1. Preparation for detection of sample nucleic acid:

[0113] 1) The inactivated sample of the new coronavirus culture is used as the positive control PC sample, and the common respiratory or lung infection pathogen sample is used as the specific interference sample for testing. These pathogen samples include Legionella pneumophila L. pneumoniae, Klebsiella pneumoniae K.pneumoniae, Streptococcus pneumoniae S.pneumoniae, Haemophilus influenzae H.influenzae, Mycoplasma pneumoniae M.pneumoniae, Chlamydia pneumoniae C.pneumoniae, B.pertussis pertussis, human coronavirus-OC43 hCoV -OC43, human coronavirus-NL63 hCoV-NL63, human coronavirus-HKU-1hCoV-HKU-1, human coronavirus-229E hCoV-229E, MERS pseudovirus MERS Armored RNA, adenovirus type 3 Adenovirus type 3, parainfluenza Type 2 Parainfluenza type 2, Respiratory Syncytial Virus A Type RSV-A, Avian Influenza Virus H7N9 Influenza A H7N9, Avian I...

Embodiment 3

[0116] Embodiment 3: the detection of the kit of the present invention to the clinical throat swab sample

[0117] 1. Preparation for detection of sample nucleic acid:

[0118] 1) Based on the throat swab samples of patients clinically diagnosed as new coronavirus infection, take 8 nucleic acid sample solutions extracted from positive throat swab samples, numbered P1, P2, P3, P4, P5, P6, P7, P8 ; 5 nucleic acid sample solutions extracted from negative throat swab samples, numbered N1, N2, N3, N4, N5, as nucleic acid samples for detection. The above throat swab nucleic acid sample solutions are all nucleic acid sample solutions obtained from clinical throat swab samples after nucleic acid extraction with the QIAamp Viral RNA Mini Kit Viral RNA Extraction Kit (Product No. 52904) from QIAGEN.

[0119] 2. The specific implementation steps of amplification and detection in this embodiment, the determination and interpretation of the detection results are the same as those in Examp...

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Abstract

The invention discloses a novel isothermal amplification CRISPR detection kit for coronavirus nucleic acid. The kit mainly comprises: 1) a high-temperature-resistant amplification enzyme; 2) an amplification buffer solution; 3) a detection enzyme; 4) a detection buffer solution; 5) mineral oil; and 6) C-tube (a reaction tube capable of separating an upper liquid reagent and a lower liquid reagent in one tube). The kit can be used for detecting the RNA of the N gene of the novel coronavirus (SARS-CoV-2) in throat swab, sputum and other samples, and has the characteristics of single tube, one step, rapidness, high specificity, high sensitivity, difficulty in pollution due to the fact that the amplification product is RNA, and the like. The kit can complete a detection reaction within about 1 hour, and the sensitivity can reach up to 3 copies per reaction or 140 copies / mL of a sample. The kit can provide molecular level judgment basis and help for clinical diagnosis of novel coronavirus infection.

Description

Technical field: [0001] The present invention relates to the molecular detection technology of novel coronavirus (SARS-CoV-2) nucleic acid, in particular to the molecular detection technology of isothermal amplification of viral RNA and CRISPR / Cas protein detection and corresponding detection kit. The invention realizes constant temperature amplification and CRISPR / Cas13a detection of novel coronavirus (SARS-CoV-2) RNA in a single tube in one step. Background technique: [0002] Coronavirus (Coronavirus) is a class of zoonotic single-stranded positive-sense RNA viruses that can be transmitted between animals and humans. At present, seven kinds of coronaviruses have been found to be able to infect humans and cause respiratory infections in humans. Human coronavirus 229E, human coronavirus OC43, human coronavirus NL63 and human coronavirus HKU1 can cause diseases with mild symptoms similar to common cold symptoms; The MERS Middle East Respiratory Syndrome that broke out in 2...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6844C12R1/93
CPCC12Q1/701C12Q1/6844C12Q2600/166C12Q2521/107C12Q2521/119C12Q2521/327C12Q2545/101C12Q2563/107C12Q2525/161C12Q2547/101Y02A50/30
Inventor 段昆付雪高晓庆施慧玲韩序王珺
Owner HANGZHOU MATRIDX BIOTECH CO LTD
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