Digital PCR, a new technique derived from traditional PCR and quantitative PCR, has revealed great potential in genetic profiling, prenatal diagnosis, cancer related allel and mutation detection. Digital PCR usually involves two steps: partitioning of limiting dilution sample into tens to thousands of individual reactions followed by PCR amplification and endpoint detection to identify the presence or absence of template molecules in each reaction. Different with real time qPCR which greatly depends on cycle threshold (C_T) and requires house-keeping gene and standard curve for relative and absolute quantification, digital PCR performs quantification through direct counting of the wells with positive amplification or calculating the concentration based on Poisson algorithm. It minimizes the measurement uncertainty caused by variation of amplification efficiency and has higher accuracy and reproducibility. In recent years, great efforts have been made to develop new methods for digital PCR to improve its sensitivity and reduce its cost. More and more techniques were commercialized soon after it was freshly published by large companies including Bio-Rad, Life technology and so on. In this article, the principle of digital PCR and its quantification method, different types of this technique and its practical applications are reviewed and discussed. Contents
1 Introduction
2 Principle of digital PCR
3 Quantification method
4 Classification of digital PCR
4.1 Microchamber/plate
4.2 Integrated fluidic circuit(IFC)
4.3 Droplet based digital PCR
5 Applications
5.1 Genetic instability analysis
5.2 Early diagnosis of cancer
5.3 Prenatal diagnosis
5.4 Others
6 Conclusion and outlook