BRAF Pyro Kit

基于序列定量检测BRAF基因中的突变

S_1084_5_GEN_V2

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BRAF Pyro Kit (24)

Cat. No. / ID:  970470

For 24 reactions: Sequencing Primers, PCR Primers, Unmethylated Control DNA, PyroMark PCR Master Mix, CoralLoad Concentrate, Buffers, and Reagents
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BRAF Pyro Kit 旨在用于分子生物学应用。该产品不能用于疾病诊断、预防和治疗。

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✓ Knowledgeable and professional Product & Technical Support

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Features

  • 实时获得全面的结果
  • 准确定量BRAF基因中的突变
  • 简便检测复杂的突变
  • 序列背景提供内置的分析参照
  • 灵活的后续分析

Product Details

BRAF Pyro Kit是用于检测BRAF基因中突变的分子检测试剂盒。该试剂盒包含扩增BRAF基因所需的引物和试剂,以及外加缓冲液、引物和试剂,用于在PyroMark Q24实时定量焦磷酸序列分析仪上采用焦磷酸测序技术实时定量检测突变。

Principle

BRAF Pyro Kit在PyroMark Q24实时定量焦磷酸序列分析仪上采用焦磷酸测序技术实时定量检测人BRAF基因中编码子600和编码子464–469的突变。BARP基因编码了v-raf鼠类肉瘤病毒癌基因homolog B1激酶,一种EGFR下游的原癌基因。研究发现,在所有癌症患者中有6–8%表现出BRAP基因的突变,在黑素瘤(50–60%)、结肠癌(10%)、甲状腺癌(39%)中BRAP基因突变则更为频繁。将近90%是V600E突变。

Procedure

该试剂盒包含2部分实验:一个用于检测编码子600的突变,另一个用于检测编码子464–469的突变(参见" Illustration of the BRAF assay")。运用PCR技术对这两个片段分别进行扩增,测定特定区域的核酸序列。使用该法进行定量和定性分析时,特定区域周围的核酸序列峰被视为标准峰和参照峰。

使用针对编码子600和编码子464–469的引物对两个片段PCR扩增后,将扩增子固定在高性能链霉素亲和琼脂糖上。将单链DNA和相应序列的引物退火处理得到DNA。然后在PyroMark Q24实时定量焦磷酸序列分析仪上设置程序和运行程序对样品进行分析(参见" Pyrogram trace of a normal genotype in codon 600"," Pyrogram trace of a normal genotype in codons 464-469"和" Pyrogram trace of a GTG to GAG mutation in base 2 of codon 600")。运行后可以通过调节"Sequence to Analyze"来检测稀有的突变。

See figures

Applications

BRAF Pyro Kit用于检测人BRAF基因中编码子600和编码子464–469的突变。

Supporting data and figures

Resources

安全数据表 (1)
Download Safety Data Sheets for QIAGEN product components.
试剂盒操作手册 (2)
For quantitative measurement of mutations in codons 600 and 464-469 of the human BRAF gene
分析软件 (1)
BRAF Plug-in 1.3
SOFTWARE (1MB)
Version 1.3.0.7
For use with PyroMark Q24 Software version 2.0.8
软件使用指南 (1)
For installation and use with PyroMark Q24 Instruments and PyroMark Q24 Software version 2.0
Safety Data Sheets (1)
Certificates of Analysis (1)
Kit Handbooks (2)
For quantitative measurement of mutations in codons 600 and 464-469 of the human BRAF gene
Analysis Software (1)
BRAF Plug-in 1.3
SOFTWARE (1MB)
Version 1.3.0.7
For use with PyroMark Q24 Software version 2.0.8
Software User Guides (1)
For installation and use with PyroMark Q24 Instruments and PyroMark Q24 Software version 2.0

FAQ

Can I use more than 10ng DNA for the PCR reaction for the Pyro kits?
The NRAS Pyro, EGFR Pyro. BRAF PyroKRAS Pyro, and UGT1A1 Pyro Kits all require that no more than 10ng of DNA be used for the PCR reaction.  Too much DNA can sometimes lead to inhibition problems in the PCR reaction.
FAQ ID -2379
What ought I do if mutation assays designed by QIAGEN fail?

Mutation assays designed  by QIAGEN detect the most common mutations. In addition, QIAGEN provides plug-ins to analyze more mutations and convenient reports for PyroMark KRAS, PyroMark EGFR, and PyroMark BRAF kits. The plug-ins can be obtained by emailing pyro.plugin@qiagen.com

 

If the mutation cannot be analyzed successfully, please send the original run files to QIAGEN Technical Service for further assistance.

FAQ ID -9065
Which purification kits are recommended for the Pyro kits?

The purification kits recommended for the BRAF Pyro, EGFR Pyro, KRAS Pyro, and NRAS Pyro kits are:

Paraffin-embedded tissue

Blood

FAQ ID -2380
What plug-ins are available for the Pyrosequencing kits?
Plug-Ins are available for:
  • EGFR Pyro Kit 
  • KRAS Pyro Kit
  • NRAS Pyro Kit
  • RAS extension Kit
  • BRAF Pyro Kit
  • These plug-ins are available for download on the respective catalog page under the Product Resources tab in the Analysis Software section. Note: PyroMark Q24 software version 2.0.7 is needed for the usage of the Plug-ins.
    FAQ ID -2381
    Is V600E the only mutation that can be detected with the codon 600 assay in the BRAF Pyro Kit?
    No, the V600E is not the only mutation that can be detected with the BRAF Pyro Kit.  The sequence to analyze can be changed post-run to allow detection also of V600M (GTG>ATG), V600A (GTG>GCG) and V600G (GTG>GGG).
    FAQ ID -2382
    Can PyroMark Gold reagents be vortexed?
    Reconstiuted enzyme and substrate of PyroMark Gold Reagents, should not be vortexed since this could lead to conformational changes which affect the activity.
    FAQ ID -2844