Comprehensive Algorithm for Quantitative Real-Time Polymerase Chain Reaction

Quantitative real-time polymerase chain reactions (qRT-PCR) have become the method of choice for rapid, sensitive, quantitative comparison of RNA transcript abundance. Useful data from this method depend on fitting data to theoretical curves that allow computation of mRNA levels. Calculating accurate mRNA levels requires important parameters such as reaction efficiency and the fractional cycle number at threshold (CT) to be used; however, many algorithms currently in use estimate these important parameters. Here we describe an objective method for quantifying qRT-PCR results using calculations based on the kinetics of individual PCR reactions without the need of the standard curve, independent of any…