用于微流控荧光PCR的高分辨率温度控制系统

范书广; 朱灵; 赵树弥; 张悦龙; 张龙; 邓国庆; 王安; 刘勇

doi:doi:10.3969/j.issn.1673-6141.2015.05.008

大气与环境光学学报, 2015, 10 (5): 417, 网络出版: 2015-10-22

用于微流控荧光PCR的高分辨率温度控制系统

High Resolution Temperature Control System of Microfluidic Fluorescence PCR

论文大纲

范书广 ¹朱灵 ^1,2,*赵树弥 ¹张悦龙 ¹张龙 ^1,2邓国庆 ¹王安 ¹刘勇 ^1,2

作者单位

¹ 中国科学院合肥物质科学研究院应用技术研究所安徽省生物医学光学仪器工程技术研究中心, 安徽合肥 230031

² 皖江新兴产业技术发展中心,安徽铜陵 244000

摘要

微流控荧光PCR产物在高分辨率熔解(HRM)曲线分析的过程中,随着DNA的解链荧光强度发生突变,而能否检测到该突变点则依赖于温控的精度。提出了多阈值PID算法,设计了一种适用于高分辨率熔解曲线分析的FPGA温度控制系统。根据测得温度值与设定值之间的差异,改变FPGA电路系统输出的脉冲宽度调制(pulse width modulation, PWM)波占空比和算法的调节频率实现对温度的准确控制。经过温度标定后,系统温度控制精度优于0.1℃,分辨率达到±0.01℃。HRM分析实验结果表明,本方法成功地辨别出了DNA序列的单碱基差异,温度控制系统达到了设计的要求。

Abstract

In the analysis of high resolution melting (HRM) of microfluidic fluorescence PCR amplification products, fluorescence intensity has a sudden change with the delink of DNA chain, while whether the abrupt change can be detected is dependent on the accuracy of temperature control. A kind of multi-threshold PID algorithm was put forward, and a temperature control system based on FPGA was designed to adapt to the analysis of high resolution melting. Based on the difference of the measurement value and the set value, the temperature change can be controlled accurately through changing the pulse-time ratio of pulse width modulation (PWM) wave and the control frequency of the algorithm from the FPGA circuit system. After temperature calibrations of the measurement system, the accuracy precedes 0.1℃ and the resolution achieves ±0.01℃. The result of HRM experiment shows that this system can detect the single base mutation of DNA sequence successfully, and the temperature control system can meet the design requirements.

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范书广, 朱灵, 赵树弥, 张悦龙, 张龙, 邓国庆, 王安, 刘勇. 用于微流控荧光PCR的高分辨率温度控制系统[J]. 大气与环境光学学报, 2015, 10(5): 417. FAN Shuguang, ZHU Ling, ZHAO Shumi, ZHANG Yuelong, ZHANG Long, DENG Guoqing, WANG An, LIU Yong. High Resolution Temperature Control System of Microfluidic Fluorescence PCR[J]. Journal of Atmospheric and Environmental Optics, 2015, 10(5): 417.

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