PCR Technology Review in GEN: Standardization of qPCR and RT-qPCR

PCR Technology Review in GEN: Standardization of qPCR and RT-qPCR
New Guidelines Seek to Promote Accurate Interpretation of Data and Reliable Results
by Stephen A. Bustin, Jo Vandesompele, Michael W. Pfaffl
published in Genetic Engineering & Biotechnology News (Vol. 29, No. 14) August 2009

The perceived ease of use of real-time quantitative PCR (qPCR) and reverse transcription PCR (RT-qPCR) technology has revolutionized life science research. Its effectiveness at amplification and quantification of low levels of nucleic acids has driven the emergence of numerous applications, including cellular mRNA and miRNA quantification, biomarker discovery and validation, microbial quantification, cancer risk assessment, gene dosage determination, and detection of extremely low copy targets for forensic investigations.This, in turn, has resulted in an abundance of publications utilizing qPCR data obtained with diverse reagents, protocols, analysis methods, and reporting formats. Unfortunately, few papers report in detail how these results were obtained. This lack of clarity and transparency has led to concern in the research community over the reliability of qPCR data interpretation and the real danger of the scientific literature being corrupted with publications reporting erroneous and conflicting results.This has already occurred in some cases, resulting, for example, in retraction of a Science “Breakthrough of the Year 2005” report. Now that qPCR has come of age, standardization is needed to ensure its validity, prompting the recent formulation of guidelines to increase experimental transparency, promote consistency between laboratories, and therefore, help assure the publication of valid conclusions.


Über Michael_W_Pfaffl