Fluorogenic DNA sequencing in PDMS microreactors.
Fluorogenic DNA sequencing in PDMS microreactors.
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DOI：
10.1038/nmeth.1629
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发表时间：
2011-06-12
期刊：
影响因子：
48
通讯作者：
Xie, Sunney
中科院分区：
文献类型：
--
作者：
Sims, Peter A.;Greenleaf, William J.;Duan, Haifeng;Xie, Sunney
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 DNA primer by DNA polymerase results in a fluorophore. We immobilize DNA templates within polydimethylsiloxane (PDMS) microreactors, sequentially introduce one of the four identically labeled TPLFNs, seal the microreactors, allow template-directed TPLFN incorporation, and measure the signal from the fluorophores trapped in the microreactors. This workflow allows sequencing in a manner akin to pyrosequencing but without constant monitoring of each microreactor. With cycle times of <10 minutes, we demonstrate 30 base reads with ∼99% raw accuracy. “Fluorogenic pyrosequencing” combines benefits of pyrosequencing, such as rapid turn-around, native DNA generation, and single-color detection, with benefits of fluorescence-based approaches, such as highly sensitive detection and simple parallelization.
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影响因子：
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作者：
Tewhey, Ryan;Warner, Jason B.;Nakano, Masakazu;Libby, Brian;Medkova, Martina;David, Patricia H.;Kotsopoulos, Steve K.;Samuels, Michael L.;Hutchison, J. Brian;Larson, Jonathan W.;Topol, Eric J.;Weiner, Michael P.;Harismendy, Olivier;Olson, Jeff;Link, Darren R.;Frazer, Kelly A.
通讯作者：
Frazer, Kelly A.
DOI：
10.1080/15257770802260741
发表时间：
2008-09
期刊：
Nucleosides, nucleotides & nucleic acids
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影响因子：
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作者：
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作者：
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