关于数字PCR的报告方昊

12数字数字 PCR(digital PCR(digital PCR,dPCRPCR,dPCR)技术是近年技术是近年来迅速发展起来的一种定量分析技术。与传统来迅速发展起来的一种定量分析技术。与传统定量定量 PCR PCR 技术不同的是数字技术不同的是数字 PCR PCR 不依赖于扩不依赖于扩增曲线的循环阈值增曲线的循环阈值(CT(CT值值)进行定量，不受扩增进行定量，不受扩增效率的影响，也不必采用看家基因和标准曲线效率的影响，也不必采用看家基因和标准曲线，是是对起始样的绝对定量。对起始样的绝对定量。3主 要 内 容数字数字 PCR 的发展与原理的发展与原理数字数字 PCR 的应用的应用312数字数字 PCR 的前景展望的前景展望420 20 世纪末，世纪末，Vogelstein Vogelstein 等提出数字等提出数字 PCRPCR 的概念，通的概念，通过将一个样本稀释后分成几十到几万份甚至上千万份，然过将一个样本稀释后分成几十到几万份甚至上千万份，然后分配到不同的反应单元，每个单元不包含或包含一个或后分配到不同的反应单元，每个单元不包含或包含一个或多个拷贝的目标分子多个拷贝的目标分子(DNA(DNA 模板模板)，在每个反应单元中分，在每个反应单元中分别对目标分子进行别对目标分子进行 PCR PCR 扩增，扩增结束后对各个反应单扩增，扩增结束后对各个反应单元的荧光信号进行统计学分析。元的荧光信号进行统计学分析。【1】56传统 PCR 或定量 PCR 反应都是在 96/384 孔板中进行的，因此早期的数字 PCR 技术也采用 96/384孔板作为反应单元。这远远不能满足数字 PCR对反应单元数的需求，于是开始制造微升和纳升级别的反应室，在提高灵敏度的同时减少了反应体系的总体积。1.微反应室/孔板数字 PCR72.微流控数字 PCR随着微流芯片技术的发展，数字 PCR 借助微流芯片，可以把样本准确并且快速的分成若干个独立的反应单元，各个单元之间互不影响，可以多步反应同时进行。提高反应通量的同时也降低了反应消耗成本。83.微滴数字 PCR微滴数字 PCR 借鉴了乳液 PCR，即将 DNA 模板与连接引物的磁性微球以极低的浓度(比如单拷贝)包裹于油水两相形成的纳升至皮升级液滴中进行 PCR 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