基于PMMA-纸基复合芯片的等离子体驱动纳米局域超快多重PCR研究

核酸检测对于疾病诊断、食品安全和环境监测至关重要，PCR是核酸检测金标准。针对传统PCR依赖于大型仪器，存在效率低、耗时长、成本高等问题。受表面等离子共振技术和复合微流控芯片平台启发，本项目拟通过等离子体驱动纳米局域热循环方法实现超快PCR扩增，通过PMMA-纸基复合微流控芯片实现多重PCR产物检测，最终开发基于PMMA-纸基复合微流控芯片的纳米局域超快多重PCR技术：通过建立光热转换与传热数值模拟并结合实验，建立纳米局域温度场的精确测量方法，探究纳米局域温度场的影响参数，实现超快速升降温（～10^4 ℃/s）及其精确控制；通过分析生物分子的热动力学，揭示纳米局域环境对多重PCR扩增的影响规律和机制；建立基于PMMA-纸基复合微流控芯片的纳米局域超快多重PCR技术用于假病毒质控品的检测。本项目研究成果将为纳米局域内光热核酸扩增技术奠定理论基础，并提供一种新的超快多重核酸即时检测工具。

Nucleic acid detection is very important for disease diagnosis, food safety and environmental monitoring, where PCR is the gold standard. Traditional PCR relies on large-scale instruments, meanwhile it is also limited by its low efficiency, long turnover time and high cost. Inspired by surface plasmon resonance (SPR) technology and hybrid microfluidic chip platform, this project intends to realize ultrafast PCR amplification by plasmon-driven nano-localized thermocycling, and realize multiplex detection of PCR product by PMMA-paper hybrid microfluidic chip, and finally develop a nano-localized ultrafast multiplex PCR technology based on PMMA-paper hybrid microfluidic chip. Through establishing numerical simulation of photothermal conversion and nano-localized heat transfer and integrating with experimental measurement of surface temperature, the accurate measurement method of nano-localized temperature field is developed. By studying the influence parameters of nano-localized temperature field, the ultrafast heating and cooling rates (~10^4 ℃/s) are realized and accurately controlled. By analyzing the thermokinetics of biomolecules, the influence and mechanism of nano-localized environment (e.g., components and their concentrations) on multiplex PCR amplification were revealed. The nano-localized ultrafast multiplex PCR technology based on PMMA-paper hybrid microfluidic chip is developed for the detection of pseudovirus quality control materials. The research results of this project will lay a theoretical foundation for the nano-localized photothermal nucleic acid amplification technology, and provide a new ultrafast multiplex tool for the point-of-care testing of nucleic acids.