极速热循环荧光定量PCR系统的设计及性能评估

Construction and performance evaluation of ultra-fast thermal cycle fluorescence quantitative PCR system

目的:设计一种用于病原体核酸现场检测的极速热循环荧光定量PCR系统,并进行性能评估。方法:该系统由扁平反应杯、极速热循环模块、固化光路荧光检测模块以及智能手机平台数据处理模块组成。极速热循环模块由升温单元和降温单元组成,其中升温单元以陶瓷片和Ag/Pb合金制成,降温单元由高速磁悬浮冷却风扇和双曲线形喉道组成;固化光路荧光检测模块采用注塑工艺制作,由光源激发单元和光探测器单元组成;智能手机平台数据处理模块包括蓝牙串口转接单元和手机应用程序2个部分,其中蓝牙串口转接单元采用TI公司的C2540F256芯片,手机应用程序由Android Studio开发。采用该系统检测甲型/乙型流感病毒和新型冠状病毒,验证该系统的检测性能。结果:该系统可实现对甲型/乙型流感病毒核酸和新型冠状病毒核酸的快速检测,且检测结果与常规实时定量PCR检测结果的一致性较好。结论:该系统体积小、质量轻,可实现病原体核酸快速现场检测。

Objective To design an ultra-fast thermal cycle fluorescent quantitative PCR system for on-site detection of pathogenic nucleic acids and evaluate its performances.Methods An ultra-fast thermal cycle fluorescent quantitative PCR system was developed with the components of a flat reaction cup,an ultra-fast thermal cycle module,a fluorescence detection module with fixed optical path and a data processing module based on the smartphone platform.The ultra-fast thermal cycle module was composed of a heating unit and a cooling unit,of which the heating unit was made of ceramic sheet and Ag/Pb alloy and the cooling unit consisted of a high-speed magnetic levitation cooling fan and a double-curved throat;the fluorescence detection module with fixed optical path was prepared with injection molding process,and made up of a light source excitation unit and a light detector unit;the data processing module based on the smartphone platform included a Bluetooth serial port adapter unit and a smartphone App,which used C2540F256 chip from TI company for developing the Bluetooth serial port adapter and Android Studio for the App.The ultra-fast thermal cycle fluorescent quantitative PCR system was used to detect influenza A/B virus and SARS-CoV-2 to verify its performances.Results The ultra-fast thermal cycle fluorescent quantitative PCR system realized rapid nucleic acid detection of influenza A/B virus and SARS-CoV-2,and the detection results were in high agreement with those by conventional real-time quantitative PCR.Conclusion The ultra-fast thermal cycle fluorescence quantitative PCR system gains advantages in small size and light weight,and can be used for rapid on-site detection of pathogen nucleic acids.