Lou Xing Jian, Panaro Nicholas J, Wilding Peter, Fortina Paolo, Kricka Larry J
University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA.
Biotechniques. 2004 Sep;37(3):392, 394, 396-8. doi: 10.2144/04373ST03.
The ligase chain reaction (LCR) following PCR is one of the most sensitive and specific methods for detecting mutations, especially single nucleotide polymorphisms (SNPs). Performing LCR in microchips remains a challenge because of the inhibitory effect of the internal surfaces of silicon-glass microchips. We have tested a dynamic polymer-based surface passivation method for LCR conducted in oxide-coated silicon-glass microchips. The combination of polyvinylpyrrolidone 40 (PVP-40) at 0.75% (w/v) with an excess of the ligase produced successful LCR in the silicon-glass microchips, with yields of ligated primers comparable to reactions performed in conventional reaction tubes. Ligated primers were detected and quantified simply and conveniently using microchip capillary electrophoresis.
聚合酶链式反应（PCR）之后的连接酶链式反应（LCR）是检测突变，尤其是单核苷酸多态性（SNP）最灵敏、最特异的方法之一。由于硅 - 玻璃微芯片内表面的抑制作用，在微芯片中进行LCR仍然是一项挑战。我们测试了一种基于动态聚合物的表面钝化方法，用于在涂有氧化物的硅 - 玻璃微芯片中进行LCR。0.75%（w/v）的聚乙烯吡咯烷酮40（PVP - 40）与过量连接酶的组合在硅 - 玻璃微芯片中成功实现了LCR，连接引物的产量与在传统反应管中进行的反应相当。使用微芯片毛细管电泳可以简单方便地检测和定量连接引物。
