报告题目：Fluorogenic DNA Sequencing in PDMS Microreactors

 

报 告 人：Dr. Peter Sims

 

Department of Chemistry and Chemical Biology，Harvard University

 

时 间：2011年4月29日星期五上午10点

 

地 点：BIOPIC 208会议室（临时入口在45乙楼东侧）

 

ABSTRACT

 

Massively parallel sequencing has emerged as a powerful analytical tool with the potential to transform medicine. We have developed a multiplex sequencing-by-synthesis method combining terminal-phosphate labeled fluorogenic nucleotides (TPLFNs) and resealable microreactors. In the presence of phosphatase, the incorporation of a non-fluorescent TPLFN into a primer strand of DNA by DNA polymerase results in the generation of a fluorophore. We immobilize DNA templates within polydimethylsiloxane (PDMS) microreactors, sequentially introduce one of the four species of TPLFN, seal the microreactors, allow template-directed TPLFN incorporation, and measure the fluorescence from each chamber. This workflow allows DNA sequencing in a manner akin to pyrosequencing but with a fluorescence signal. “Fluorogenic pyrosequencing” combines benefits of pyrosequencing, such as rapid turn-around times, generation of native DNA, and single-color detection, with benefits of fluorescence-based approaches, such as highly sensitive detection and simple parallelization.