Surface-enhanced Raman scattering based ligase detection reaction.
Surface-enhanced Raman scattering based ligase detection reaction.
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DOI：
10.1021/ja807526v
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发表时间：
2009-02-18
影响因子：
15
通讯作者：
Erickson D
中科院分区：
文献类型：
--
作者：
Huh YS;Lowe AJ;Strickland AD;Batt CA;Erickson D
基因组学提供了生物体完整基因组成的全面视图。单核苷酸多态性 (SNP) 所体现的个体序列变异可以深入了解包括癌症在内的大量表型和疾病的基础。快速筛选 SNP 的能力将对许多应用产生深远的影响，尤其是个性化医疗。在这里，我们展示了一种通过将表面增强拉曼散射 (SERS) 应用于连接酶检测反应 (LDR) 进行 SNP 检测的新方法。该反应使用两种 LDR 引物，其中一种含有拉曼增强剂，另一种含有报告染料。在 LDR 中，其中一个引物被设计用于询问 SNP。当被询问的 SNP 与鉴别引物序列相匹配时，引物就会被连接，增强子和染料就会非常接近，从而能够检测到染料的拉曼特征。通过检测染料的拉曼特征而不是其荧光发射，我们的技术避免了光谱重叠的问题，该问题限制了现有系统可以并行进行的反应数量。我们展示了用于检测人类 K-ras 癌基因点突变的 LDR-SERS 反应。该反应在电动活性微流体装置中实现，该装置能够对反应产物进行物理浓缩，以增强检测灵敏度和量化。我们报告的目标 DNA 检测限为 20 pM，具有 LDR 平台产生的预期特异性。
Genomics provides a comprehensive view of the complete genetic makeup of an organism. Individual sequence variations, as manifested by single nucleotide polymorphisms (SNPs), can provide insight into the basis for a large number of phenotypes and diseases including cancer. The ability rapidly screen for SNPs will have a profound impact on a number of applications, most notably personalized medicine. Here we demonstrate a new approach to SNP detection through the application of surface-enhanced Raman scattering (SERS) to the ligase detection reaction (LDR). The reaction uses two LDR primers, one of which contains a Raman enhancer and the other a reporter dye. In LDR, one of the primers is designed to interrogate the SNP. When the SNP being interrogated matches the discriminating primer sequence, the primers are ligated and the enhancer and dye are brought into close proximity enabling the dye’s Raman signature to be detected. By detecting the Raman signature of the dye rather than its fluorescence emission, our technique avoids the problem of spectral overlap which limits number of reactions which can be carried out in parallel by existing systems. We demonstrate the LDR-SERS reaction for the detection of point mutations in the human K-ras oncogene. The reaction is implemented in an electrokinetically active microfluidic device that enables physical concentration of the reaction products for enhanced detection sensitivity and quantization. We report a limit of detection of 20 pM of target DNA with the anticipated specificity engendered by the LDR platform.
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影响因子：
5.1
作者：
Nakitandwe J;Trognitz F;Trognitz B
通讯作者：
Trognitz B
影响因子：
12.6
作者：
Fang, Cheng;Agarwal, Ajay;Kwong, Dim-Lee
通讯作者：
Kwong, Dim-Lee
影响因子：
6.2
作者：
Jung, Hye Young;Park, Yong-Kyun;Kim, Seong Kyu
通讯作者：
Kim, Seong Kyu
影响因子：
3.3
作者：
Doering, WE;Nie, SM
通讯作者：
Nie, SM
影响因子：
15
作者：
Fabris, Laura;Dante, Mark;Bazan, Guillermo C.
通讯作者：
Bazan, Guillermo C.