超快脉冲控制PCR（upPCR）技术及应用

在精准医疗、个性化医疗的大背景下，分子诊断在病原体检测、肿瘤诊断、优生优育、环境保护、食品安全等领域的应用越来越广泛，并逐渐向操作简单、快速准确、低成本、适用于基层及家庭使用的分子即时检测（point-of-care testing，POCT）方向发展。超快脉冲控制PCR（ultra-fast pulse-controlled PCR，upPCR）是实时荧光定量PCR（qPCR）技术的延伸和升级，该技术利用能量脉冲控制扩增反应中的金属加热元件（主要是纳米金），在几百微秒内完成溶液局部微环境的快速升温，实现模板DNA的解链变性，停止加热后反应微环境可被周围溶液快速冷却到聚合酶的延伸温度，实现引物退火和模板DNA的扩增，单个变性-扩增循环仅有1.5~5 s，远快于传统PCR（约90 s/循环），从而能够极大地加快扩增反应速度。upPCR技术在保留了传统qPCR高灵敏度、高特异性和多重检测等优势的基础上，增加了超快速（低于15 min）、设备简单等新优势，非常适合用于基层检测等分子POCT场景。本文主要对upPCR技术的原理、设备、核心原料及在分子诊断中的应用进行综述，并对该技术存在的优缺点，以及未来的技术发展和应用趋势进行了讨论。

Applications of Ultra-fast Pulse-controlled PCR (upPCR)

Under the background of precision medicine and personalized medicine, molecular diagnosis is more and more widely applied in pathogen detection, tumor diagnosis, eugenics and fertility, environmental protection, food safety and other fields, and continues to develop in the direction of molecular point-of-care testing (POCT) with advantages of fast and accurate, low-cost, simple operation. Ultra-fast pulse-controlled PCR (upPCR) is an extension and upgrade of real-time quantitative PCR (qPCR) technology, which uses energy pulses to control metal heating elements (mainly gold nanoparticles) in amplification reactions to complete the rapid heating of the local microenvironment of the solution within a few hundred microseconds, and realize the melting and denaturation of template DNA; after stopping heating, the reaction microenvironment can be rapidly cooled by the surrounding solution down to the extension temperature of the polymerase to achieve amplification of template DNA. A single denaturation-amplification cycle of upPCR is only 1.5-5 s, which is much faster than traditional PCR (about 90 s per cycle), and can thus greatly speed up the PCR reaction. On the basis of retaining the advantages of traditional qPCR such as high sensitivity, high specificity and multiplex detection, ultra-fast pulse control PCR technology adds new advantages such as ultra-fast reaction time (less than 15 min) and simple operation, which is very suitable for molecular POCT scenarios such as home detection and community screening. This paper mainly reviews the principle, core raw materials, equipments and applications of upPCR technology in molecular diagnosis, and discusses the advantages and disadvantages of this technology, as well as future technology development and application trends.