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Development of high sensitive and high S/N ratio multicolored SNP detection technology toward each locus.

开发针对每个位点的高灵敏度、高信噪比的多彩SNP检测技术。

基本信息

批准号：
20300183
负责人：
LEZHAVA Alexander
金额：
$ 12.56万
依托单位：
The Institute of Physical and Chemical Research
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2008
资助国家：
日本
起止时间：
2008 至 2010
项目状态：
已结题

项目摘要

A two-color readout of the Smart Amplification (SmartAmp、SMAP) genotyping system using so-called Exciton Primers have been developed. New method is highly specific and sensitive, permitting a one-step, single-tube mutation detection reaction which delivers a result in 45 minutes directly from blood. Exciton Primers, which are functioning as "fluorescence emission with sequence-specific," after hybridization to complementary sequences, can provide the signals resulting target sequence detection for real-time monitoring of amplification reactions. Applied to the isothermal SmartAmp2 mutation detection process, Exciton Primers show high signal strength with low background leading to a superior specificity and sensitivity compared to SYBR Green I. The genotyping assay can use only one labeled Exciton Primer for endpoint detection, or simultaneously by real-time monitoring detect wild-type and mutant alleles in a one-tube reaction using two Exciton Primers having different dyes. The technique seems very promising for decentralized point-of-care diagnostics since the approach does not involve complicated instrumentation and sample purification. We demonstrated efficiency of the new technology on the example of rapid detection of SNPs and mutations

使用所谓的激子引物的智能扩增（SmartAmp、SMAP）基因分型系统的双色读数已经开发出来。新方法具有高度特异性和敏感性，可实现一步式单管突变检测反应，可在 45 分钟内直接从血液中得出结果。激子引物在与互补序列杂交后起到“序列特异性荧光发射”的作用，可以提供目标序列检测的信号，用于实时监测扩增反应。应用于等温SmartAmp2突变检测过程中，Exciton Primers显示出高信号强度和低背景，与SYBR Green I相比具有优异的特异性和灵敏度。基因分型测定可以仅使用一种标记的Exciton Primer进行终点检测，或者使用两种具有不同染料的Exciton Primers在一管反应中同时通过实时监测检测野生型和突变型等位基因。该技术对于分散式护理诊断来说似乎非常有前景，因为该方法不涉及复杂的仪器和样品纯化。我们通过快速检测 SNP 和突变的例子证明了新技术的效率