Wang Lin, Li Paul C H, Yu Hua-Zhong, Parameswaran Ash M
Department of Chemistry, Simon Fraser University, 8888 University Drive, Burnaby, V5A 1S6, BC, Canada.
Anal Chim Acta. 2008 Mar 3;610(1):97-104. doi: 10.1016/j.aca.2007.12.048. Epub 2008 Jan 17.
Detection of polymerase chain reaction (PCR) products obtained from cultured greenhouse fungal pathogens, Botrytis cinerea and Didymella bryoniae has been achieved using a previously developed microfluidic microarray assembly (MMA) device. The flexible probe construction and rapid DNA detection resulted from the use of centrifugal pumping in the steps of probe introduction and sample delivery, respectively. The line arrays of the oligonucleotide probes were "printed" on a CD-like glass chip using a polydimethylsiloxane (PDMS) polymer plate with radial microfluidic channels, and the sample hybridizations were conducted within the spiral channels on the second plate. The experimental conditions of probe immobilization and sample hybridization were optimized, and both complementary oligonucleotides and PCR products were tested. We were able to achieve adequate fluorescent signals with a sample load as small as 0.5 nM (1 microL) for oligonucleotide samples; for PCR products, we achieved detection at the level of 3 ng.
利用先前开发的微流控微阵列组件（MMA）装置，已实现对从温室培养的真菌病原体灰葡萄孢菌和黄瓜壳二孢中获得的聚合酶链反应（PCR）产物的检测。在探针引入和样品输送步骤中分别使用离心泵送，实现了灵活的探针构建和快速的DNA检测。寡核苷酸探针的线性阵列使用带有径向微流控通道的聚二甲基硅氧烷（PDMS）聚合物板“印刷”在类似CD的玻璃芯片上，样品杂交在第二块板上的螺旋通道内进行。对探针固定和样品杂交的实验条件进行了优化，并对互补寡核苷酸和PCR产物进行了测试。对于寡核苷酸样品，我们能够在低至0.5 nM（1微升）的样品负载量下获得足够的荧光信号；对于PCR产物，我们实现了3 ng水平的检测。
