Development of Real-Time Polymerase Chain Reaction on a Chip by Bottom Heating
Date Issued
2009
Date
2009
Author(s)
Hou, Chien-Cheng
Abstract
The concept of Polymerase Chain Reaction primarily uses those three processes (Denaturizing, Annealing and Extension) as one cycle, and DNA will clone by the increase rate of 2n. Real-Time PCR is the most famous in the concept. The technique mainly needs to combine Thermal Control Cycler with Fluorimeter, and do DNA outside-body proliferation in thermal control process. And meanwhile, in order to monitor Labeling Dye in proliferation process, the change of fluorescence level and do quantitative analysis, the instrument called Real-time PCR machine is needed to achieve the above-mentioned technique. So Thermal Cycler and Fluorimeter are the keys to quantitative analysis.he first part of the thesis will discuss the difficulty that Lab-on-a-Chip is heated-from-the bottom system. When Heated-from-the bottom system controls the bottom heated temperature to do the Polymerase Chain Reaction process, it is possible to make inner reagent cause the phenomena of temperature gradient and natural convection. When Using thermocouple measures the bottom temperature of Lab-on-a -Chip and upper temperature of reagent, we find there is a gradient temperature up to 9.7℃.In order to solve the problem, changing reagent volume or adding DMSO to reagent are proposed.he second part of the thesis will discuss how the fluorescence detection system operate in DNA fluorescence detection. In the study, we use 350bp(base pair) Hepatitis B virus, with 108 copies/ml DNA template as experiment reagent, and besides, adding SyBr Green dye for fluorescence detection. Because fluorescence signal is invisible to the naked eye, we use MATLAB to cope with the fluorescence signals filmed by CCD to quantify them. Thus we can get weak DNA fluorescence signals which are used for quantitative analysis.
Subjects
PCR
DMSO
HBV
Rayleigh Number
SDGs
Type
thesis
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