High-throughput droplet digital PCR system for absolute quantitation of DNA copy number.
High-throughput droplet digital PCR system for absolute quantitation of DNA copy number.
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DOI：
10.1021/ac202028g
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发表时间：
2011-11-15
影响因子：
7.4
通讯作者：
Colston, Bill W.
中科院分区：
文献类型：
--
作者：
Hindson, Benjamin J.;Ness, Kevin D.;Masquelier, Donald A.;Belgrader, Phillip;Heredia, Nicholas J.;Makarewicz, Anthony J.;Bright, Isaac J.;Lucero, Michael Y.;Hiddessen, Amy L.;Legler, Tina C.;Kitano, Tyler K.;Hodel, Michael R.;Petersen, Jonathan F.;Wyatt, Paul W.;Steenblock, Erin R.;Shah, Pallavi H.;Bousse, Luc J.;Troup, Camille B.;Mellen, Jeffrey C.;Wittmann, Dean K.;Erndt, Nicholas G.;Cauley, Thomas H.;Koehler, Ryan T.;So, Austin P.;Dube, Simant;Rose, Klint A.;Montesclaros, Luz;Wang, Shenglong;Stumbo, David P.;Hodges, Shawn P.;Romine, Steven;Milanovich, Fred P.;White, Helen E.;Regan, John F.;Karlin-Neumann, George A.;Hindson, Christopher M.;Saxonov, Serge;Colston, Bill W.
数字PCR可以在样本中绝对定量核酸。使用96孔板工作流程的常规Taqman分析，可以处理约200万个PCR反应。我们提供的DDPCR系统提供的数量级比实时PCR更精确和灵敏度，我们显示了生殖线拷贝数的精度。折叠超过野生型背景，我们证明了对胎儿的绝对定量和无细胞等离子体的母体DNA。探索复杂的遗传景观，发现和验证新的疾病关联，并定义分子诊断的新时代。
Digital PCR enables the absolute quantitation of nucleic acids in a sample. The lack of scalable and practical technologies for digital PCR implementation has hampered the widespread adoption of this inherently powerful technique. Here we describe a high-throughput droplet digital PCR (ddPCR) system that enables processing of ∼2 million PCR reactions using conventional TaqMan assays with a 96-well plate workflow. Three applications demonstrate that the massive partitioning afforded by our ddPCR system provides orders of magnitude more precision and sensitivity than real-time PCR. First, we show the accurate measurement of germline copy number variation. Second, for rare alleles, we show sensitive detection of mutant DNA in a 100 000-fold excess of wildtype background. Third, we demonstrate absolute quantitation of circulating fetal and maternal DNA from cell-free plasma. We anticipate this ddPCR system will allow researchers to explore complex genetic landscapes, discover and validate new disease associations, and define a new era of molecular diagnostics.
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影响因子：
30.8
作者：
Alkan, Can;Kidd, Jeffrey M.;Marques-Bonet, Tomas;Aksay, Gozde;Antonacci, Francesca;Hormozdiari, Fereydoun;Kitzman, Jacob O.;Baker, Carl;Malig, Maika;Mutlu, Onur;Sahinalp, S. Cenk;Gibbs, Richard A.;Eichler, Evan E.
通讯作者：
Eichler, Evan E.
影响因子：
6
作者：
Kauraniemi, P;Kuukasjärvi, T;Kallioniemi, A
通讯作者：
Kallioniemi, A
影响因子：
56.9
作者：
Gonzalez, E;Kulkarni, H;Ahuja, SK
通讯作者：
Ahuja, SK
影响因子：
3.7
作者：
Dube S;Qin J;Ramakrishnan R
通讯作者：
Ramakrishnan R
影响因子：
9.8
作者：
Fan, H. Christina;Blumenfeld, Yair J.;El-Sayed, Yasser Y.;Chueh, Jane;Quake, Stephen R.
通讯作者：
Quake, Stephen R.