Xu Lulu, Duan Jiaxin, Chen Junman, Ding Shijia, Cheng Wei
The Center for Clinical Molecular Medical Detection, The First Affiliated Hospital of Chongqing Medical University, Chongqing, 400016, PR China.
Key Laboratory of Clinical Laboratory Diagnostics (Ministry of Education), College of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016, PR China.
Anal Chim Acta. 2021 Mar 1;1148:238187. doi: 10.1016/j.aca.2020.12.062. Epub 2020 Dec 31.
Rolling circle amplification (RCA) is an efficient enzymatic isothermal reaction that using circular probe as a template to generate long tandem single-stranded DNA or RNA products under the initiation of short DNA or RNA primers. As a simplified derivative of natural rolling circle replication which synthesizes copies of circular nucleic acids molecules such as plasmids, RCA amplifies the circular template rapidly without thermal cycling and finds various applications in molecular biology. Compared with other amplification strategies, RCA has many obvious advantages. Firstly, because of the strict complementarity required in ligation of a padlock probe, it endows the RCA reaction with high specificity and can even be utilized to distinguish single base mismatches. Secondly, through the introduction of multiple primers, exponential amplification can be achieved easily and leads to a good sensitivity. Thirdly, RCA products can be customized by manipulating circular templates to generate functional nucleic acids such as aptamer, DNAzymes and restriction enzyme sites. Moreover, the RCA has good biocompatibility and is especially suitable for in situ detection. Therefore, RCA has attracted considerable attention as an efficient and potential tool for highly sensitive detection of biomarkers. Herein, we comprehensively introduce the fundamental principles of RCA technology, summarize it from three aspects including initiation mode, amplification mode and signal output mode, and discuss the recent application of RCA-based biosensor in this review.
滚环扩增（RCA）是一种高效的酶促等温反应，它以环状探针为模板，在短DNA或RNA引物的引发下生成长串联单链DNA或RNA产物。作为自然滚环复制的简化衍生物，自然滚环复制用于合成环状核酸分子（如质粒）的拷贝，RCA无需热循环即可快速扩增环状模板，并在分子生物学中有多种应用。与其他扩增策略相比，RCA有许多明显的优点。首先，由于锁式探针连接需要严格互补，这赋予RCA反应高特异性，甚至可用于区分单碱基错配。其次，通过引入多个引物，可以轻松实现指数扩增并具有良好的灵敏度。第三，RCA产物可以通过操纵环状模板进行定制，以生成功能性核酸，如适体、脱氧核酶和限制性酶切位点。此外，RCA具有良好的生物相容性，特别适合原位检测。因此，RCA作为一种用于生物标志物高灵敏度检测的高效且有潜力的工具，已引起了相当大的关注。在此，我们全面介绍RCA技术的基本原理，从引发模式、扩增模式和信号输出模式三个方面对其进行总结，并在本综述中讨论基于RCA的生物传感器的最新应用。
