Statistical Analysis of Quantitative PCR Data

Abstract

This thesis seeks to develop a better understanding of the analysis of gene expression to find the amount of transcript in a sample. The mainstream method used is called Polymerase Chain Reaction (PCR) and it exploits the DNA's ability to replicate. The comparative CT method estimate the starting fluorescence level f0 by assuming constant amplification in each PCR cycle, and it uses the fluorescence level which has risen above a certain threshold. We present a generalization of this method, where different threshold values can be used. The main aim of this thesis is to evaluate a new method called the Enzymological method. It estimates f0 by considering a cycle dependent amplification and uses a larger part of the fluorescence curves, than the two CT methods. All methods are tested on dilution series, where the dilution factors are known. In one of the datasets studied, the Clusterin dilution-dataset, we get better estimates from the Enzymological method compared to the two CT methods.