EGFR Pyro Kit

基于序列定量分析EGFR基因中的突变

S_1084_5_GEN_V2

✓ 24/7 automatic processing of online orders

✓ Knowledgeable and professional Product & Technical Support

✓ Fast and reliable (re)-ordering

EGFR Pyro Kit (24)

Cat. No. / ID:  970480

For 24 reactions: Sequencing Primers, PCR Primers, Unmethylated Control DNA, PyroMark PCR Master Mix, CoralLoad Concentrate, Buffers, and Reagents
Copy order details
Log in To see your account pricing.
EGFR 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。

✓ 24/7 automatic processing of online orders

✓ Knowledgeable and professional Product & Technical Support

✓ Fast and reliable (re)-ordering

Features

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

Product Details

EGFR Pyro Kit是用于检测EGFR基因中突变和缺失的分子检测试剂盒。该试剂盒包含扩增EGFR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

EGFR Pyro Kit用于实时定量检测人EGFR基因编码子719、768、790、858和861的突变,以及外显子19的缺失和复杂突变,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进行分析。EGFR基因可编码表皮生长因子受体(EGFR)蛋白。EGFR基因酪氨酸激酶结构域的突变能够促进肿瘤的生长和增殖。在美国有近10%的非小细胞肺癌患者发现了EGFR基因突变,在东亚该比例达到35%。除此之外,6%的脑瘤患者也发现了EGFR突变。

可检测以下种类的突变:

  • 外显子18(719):G719A、G719C和G719S
  • 外显子19(缺失):20种缺失和复杂突变
  • 外显子20(768和790):S768I和T790M
  • 外显子21(858–861):L858R、L861Q和L861R 

Procedure

该试剂盒包含4个PCR实验:第一个用于检测编码子719的突变(外显子18);第二个用于检测编码子768和790的突变(外显子20);第三个用于检测编码子858–861的突变(外显子21);第四个用于鉴定外显子19的缺失突变。对这4个片段分别进行PCR扩增,然后测定特定区域的核酸序列(参见" Illustration of the EGFR assay")。其中包含编码子768和790的扩增子被分为两个序列反应。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。

使用针对外显子18、19、20和21的引物对4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。推荐使用EGFR Plug-in Report进行分析。该报告确保使用正确的LODs和分析的不同序列可自动检测所有突变和缺失。也可在PyroMark Q24实时定量焦磷酸序列分析仪上使用其他分析工具进行分析(参见" Pyrogram trace of a normal genotype in codon 719"," Pyrogram trace of a normal genotype in codon 768"," Pyrogram trace of a normal genotype in exon 19"和" Pyrogram trace of a GGAATTAAGAGAAGC deletion in exon 19")。运行后可以通过调节"Sequence to Analyze"来检测外显子19的各种缺失和其他外显子的稀有突变。

See figures

Applications

EGFR Pyro Kit用于检测人EGFR基因编码子719、768、790、858和861的突变,以及外显子19的缺失和复杂突变。

可检测以下种类的突变:

  • 外显子18(719):G719A、G719C和G719S
  • 外显子19(缺失):20种缺失和复杂突变
  • 外显子20(768和790):S768I和T790M
  • 外显子21(858–861):L858R、L861Q和L861R

Supporting data and figures

Resources

软件使用指南 (1)
For installation and use with PyroMark Q24 Instruments and PyroMark Q24 Software version 2.0
安全数据表 (1)
Download Safety Data Sheets for QIAGEN product components.
分析软件 (1)
EGFR Plug-in 1.3
SOFTWARE (2MB)
Version 1.3.0.7
For use with PyroMark Q24 Software version 2.0.8
Safety Data Sheets (1)
Certificates of Analysis (1)
Analysis Software (1)
EGFR Plug-in 1.3
SOFTWARE (2MB)
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
    Can the insertions in Exon 20 be detected with the EGFR Pyro Kit?
    No, there is no specific assay for the insertions and the codon 768 assay does not allow detection of the insertions starting at amino acid 770.
    FAQ ID -2383
    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