GMO Detection Laboratory, SJTU-Bor Luh Food Safety Center, School of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, People's Republic of China.
Anal Chem. 2011 Mar 1;83(5):1579-86. doi: 10.1021/ac103266w. Epub 2011 Feb 3.
We describe the development of a novel combined approach for high-throughput analysis of multiple DNA targets based on multiplex Microdroplet PCR Implemented Capillary gel electrophoresis (MPIC), a two-step PCR amplification strategy. In the first step, the multiple target DNAs are preamplified using bipartite primers attached with universal tail sequences on their 5'-ends. Then, the preamplified templates are compartmentalized individually in the microdroplet of the PCR system, and multiple targets can be amplified in parallel, employing primers targeting their universal sequences. Subsequently, the resulting multiple products are analyzed by capillary gel electrophoresis (CGE). Using genetically modified organism (GMO) analysis as a model, 24 DNA targets can be simultaneously detected with a relative limit of detection of 0.1% (w/w) and absolute limit of detection of 39 target DNA copies. The described system provides a promising alternative for high-throughput analysis of multiple DNA targets.
我们描述了一种新的基于多重微滴 PCR 实施毛细管凝胶电泳（MPIC）的高通量分析多种 DNA 靶标的联合方法的开发，这是一种两步 PCR 扩增策略。在第一步中，使用带有通用尾部序列的双部分引物对多个目标 DNA 进行预扩增。然后，将预扩增模板单独分区到 PCR 系统的微滴中，并且可以使用针对其通用序列的引物来平行扩增多个目标。随后，通过毛细管凝胶电泳（CGE）分析所得的多个产物。使用转基因生物（GMO）分析作为模型，可同时检测 24 个 DNA 靶标，相对检测限为 0.1%（w/w），绝对检测限为 39 个目标 DNA 拷贝。所描述的系统为高通量分析多种 DNA 靶标提供了一种有前途的替代方法。
