Research On The Temperature Control And Fluorescence Signal Processing Of Micro-droplet Digital PCR System

Polymerase chain reaction(PCR)is a technique used in molecular biology to amplify a single copy or a few copies of a segment of DNA across several orders of magnitude,generating thousands to millions of copies of a particular DNA sequence.Digital PCR instrument(DPCRI)can well amplify and quantify target DNA.Therefore,DPCRI has attracted great attentions and been widely applied.This thesis firstly reviews the state-of-the-art of DPCRI.Based on the review,it investigates key techniques of DPCRI,including temperature control during PCR amplification and peak-value detection of micro-droplet fluorescence signal.The main contents of this thesis are as follows:(1)It firstly analyzed the temperature requirement of PCR amplification.Based on the analyses,hardware design of DPCRI considering temperature control is developed,in detail,it integrates a heating/cooling circuit and a temperature measurement circuit,as well as a minimum-system design of CPU.Therein,the heating/cooling circuit can well satisfy the requirement of temperature related to the speed of raise and fall by selecting the type and parameter of thermoelectric cooler;while the temperature measuring circuit chooses PT100 as the temperature sensor to realize accurate feedback.It should be mentioned that fuzzy PID control algorithm is applied into the temperature control.Simulation results validate that the performance of fuzzy PID control algorithm,especially speed and precision,is superior to ones under traditional PID control.(2)It demonstrates the structure of the micro-droplet fluorescence detection system.The photoelectric detection circuit is designed,including design of photo-electric signal conversion circuit,and amplifier circuits of both former and medium stages.(3)It develops the processing procedure of micro-droplet fluorescence,including baseline adjustment,filtering,and peak-value detection.For section of baseline adjustment,it chooses sectioning method for baseline adjustment and it gets a good result.For section of filtering,FIR filtering,IIR filtering,S-G filtering are compared since the noise accompany with the acquisition of the fluorescence,and the simulation result shows that S-G filtering is better than the other two,especially in fidelity and de-noising ability.For the facticity of peak-value detection,a new peak detection based on the Gaussian strategy is applied.Simulation result shows that the new method can seek not only the peak-position accurately,but also the overlapping peak,besides,the method can also achieve a good result without filtering.This thesis studies the temperature control module and the fluorescence detection module of DPCRI,including the design of circuit,and the suitable filtering method.In addition,a new peak-value detection method is proposed.It is concluded that the design and proposed method all well agree the analyses,which verifies the effectiveness of the thesis' contributions.