Novel beta coronavirus variation detection method, probe and kit

A coronavirus, mutation detection technology, applied in biochemical equipment and methods, microbial determination/inspection, resistance to vector-borne diseases, etc., can solve problems such as large amount of sequencing data, high sequencing cost, difficult virus gene sequence detection, etc. , to achieve the effect of improving the detection sensitivity

Active Publication Date: 2020-07-24
广西识远医学检验实验室有限公司
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Problems solved by technology

However, the amount of sequencing data required by this method is huge, and the cost of sequencing is high; and this method is aimed at samples from patients in the occult period or samples with low virus abu

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  • Novel beta coronavirus variation detection method, probe and kit
  • Novel beta coronavirus variation detection method, probe and kit
  • Novel beta coronavirus variation detection method, probe and kit

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[0035] Example

[0036] 1. Materials and methods

[0037] 1.1 Reagent

[0038] 1) Viral RNA extraction kit, purchased from Tiangen Biochemical (Beijing) Technology Co., Ltd., DP315-R;

[0039] 2) RNA library construction kit, purchased from ABclonal, RK20301;

[0040] 3) The probe hybridization capture kit uses Aiji Taikang (Beijing) Biotechnology Co., Ltd. TargetSeq Onekit.

[0041] 1.2 method

[0042] 1.2.1 Probe design

[0043] Based on the published whole genome sequence of the new coronavirus and the related coronavirus genus sequences in the NCBI database, a liquid-phase probe detection kit was designed and synthesized to cover all currently known β-coronavirus whole genome sequences. The virus genome used to design the probe is as follows: 1) SARS-CoV-2: contains 32 sequences (source: 18 from Wuhan, 6 from Guangzhou, 2 from Zhejiang, 2 from the United States, 2 from Nonthaburi, Thailand, Japan 1 Kanagawa, and bat RaTG13). 2) Other β-coronavirus strains: 1122 in total. Covers SAR...

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Abstract

The invention relates to a novel beta coronavirus (SARS-CoV-2) variation detection kit. The kit is characterized by comprising the following parts: an RNA probe group, primers, a blocking buffer, an RNA enzyme blocking solution, a hybridization buffer, a binding buffer, an SSC rinsing solution with 1 time concentration, an SSC rinsing solution with 0.1 time concentration, a PCR reaction solution and a TE buffer, wherein the RNA probe group comprises 311 RNA probes, and the sequences of the RNA probes are shown as SEQ ID NO.1 to SEQ ID NO.311. The novel beta coronavirus (SARS-CoV-2) variation detection kit provided by the invention can enrich beta coronavirus nucleic acid sequences in a library, improves the detection sensitivity, detects beta coronaviruses of all people and animals at thesame time, and is used for beta coronavirus detection and hereditary variation detection.

Description

technical field [0001] The invention relates to the field of biological gene detection, in particular to a novel betacoronavirus (SARS-CoV-2) mutation detection method, probe and kit based on liquid-phase probe hybridization capture. Background technique [0002] SARS-CoV-2 and SARS coronavirus and "Middle East Respiratory Syndrome" MERS coronavirus belong to the genus βcoronavirus, and the genome is linear positive-strand RNA. As an RNA virus, the new coronavirus has a high mutation frequency, and it will mutate more during the transmission between wild animals-intermediate hosts-humans. The 2019-nCoV nucleic acid detection kit based on the real-time quantitative PCR (RT-qPCR) method only conducts qualitative detection for the specific conserved sequence region of the 2019-nCoV or the antigen of a certain strain initially released. This method has low tolerance for genetic variation. If the virus gene mutation occurs in the specific binding region of the primer and probe,...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6869C12N15/11
CPCC12Q1/701C12Q1/6869C12Q2531/113C12Q2527/125C12Q2527/127C12Q2535/122Y02A50/30
Inventor 杭兴宜孙永乐梁加龙屈武斌王瑞超蔡万世
Owner 广西识远医学检验实验室有限公司
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