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Nucleic acid composition and kit for detecting BRAF gene mutation and detection method of BRAF gene mutation

A nucleic acid composition and detection method technology are applied in the field of nucleic acid compositions for detecting BRAF gene mutation, and can solve the problems of low detection sensitivity and inability to meet actual needs and the like

Pending Publication Date: 2020-12-04
SUZHOU ZHONGKE ADVANCED TECH RES INST CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The detection sensitivity of these methods is low and cannot meet the actual needs

Method used

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  • Nucleic acid composition and kit for detecting BRAF gene mutation and detection method of BRAF gene mutation
  • Nucleic acid composition and kit for detecting BRAF gene mutation and detection method of BRAF gene mutation
  • Nucleic acid composition and kit for detecting BRAF gene mutation and detection method of BRAF gene mutation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] Provided is a kit for detecting BRAF gene mutation.

[0056] The kit includes nucleic acid composition for detecting BRAF gene mutation (as shown in Table 1), dNTPs, PCR reaction solution, enzyme mixture, positive control substance and blank control substance.

[0057] Wherein, the enzyme mixed solution includes the enzyme I mixed solution. Enzyme I mixture is a mixture of Taq enzymes and antibodies without 5' to 3' exonuclease activity. Enzyme II mixture is a mixture of exonuclease activity and antibody from 5' end to 3' end. Among them, the Taq enzyme was purchased from TAKARA Biological Company and Guangzhou Baiwang Biotechnology Co., Ltd., and the antibody was purchased from TOYOBO Company.

[0058] Mutation detection PCR reaction solution includes: 2mM ~ 5mM MgCl 2 , 20mM~50mM Tris, pH8.3, 200mM~500mg / L BSA, 50μM~100μM dNTPs, 0.2U-1U enzyme I mixture, 0.1μM~0.5μM first forward primer, 0.1μM ~0.5 μM first reverse primer, 0.1 μM ~ 0.5 μM first probe, 0.1 μM ~ 0.5...

Embodiment 2

[0065] Measure the sensitivity of the kit for detecting BRAF gene mutation of embodiment 1

[0066] Take human BRAF gene wild-type genome, human BRAF gene V600E mutant plasmid and TE buffer buffer, prepare human BRAF wild-type DNA containing 5ng / μL and V600E whose content is 10%, 1% and 0.1% of wild type respectively For mutant plasmids, record the positive detection rate.

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Abstract

The invention relates to a nucleic acid composition and kit for detecting BRAF gene mutation and a detection method of BRAF gene mutation. The nucleic acid composition for detecting BRAF gene mutationcomprises a first primer pair and a first probe, wherein the sequences of the first primer pair are shown as SEQ ID No. 1 and SEQ ID No. 2, the sequence of the first probe is shown as SEQ ID No. 3, and the first primer pair and the first probe are used for detecting V600E mutation of a BRAF gene. The specificity of the first primer pair and the first probe in the nucleic acid composition on V600Emutation of the BRAF gene is relatively high, the V600E mutation of the BRAF gene can be detected, and the detection sensitivity is relatively high.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a nucleic acid composition and kit for detecting BRAF gene mutations and a detection method for BRAF gene mutations. Background technique [0002] The BRAF gene encodes a serine / threonine-specific kinase, which is an important transduction factor in the MEK-ERK signaling pathway and participates in the regulation of various physiological processes such as cell growth, differentiation and apoptosis. Mutations in BRAF can affect various physiological processes such as cell growth, differentiation and apoptosis. The mutation form of BRAF is mainly V600E mutation (that is, the amino acid at position 600 of exon 15 of the BRAF gene is mutated from valine to glutamic acid). Therefore, the determination of the V600E mutation of the BRAF gene is of great significance. [0003] At present, there are mainly the following methods for BRAF gene V600E mutation detection: (1) Sanger sequen...

Claims

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Application Information

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IPC IPC(8): C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q2535/137C12Q2531/113C12Q2563/107C12Q2545/101
Inventor 张炳为辜嘉周树民
Owner SUZHOU ZHONGKE ADVANCED TECH RES INST CO LTD
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