Digital PCR is an absolute DNA quantification technique to determine the copy number of target DNA. PCR reaction solution is divided into aliquot throughout numerous partitions and amplifies independently. The copy number of target DNA is estimated by the statistical analysis of positive signals. Random and independent distribution of the target DNA molecules throughout the partitions of the digital panel and successful amplification from single molecules are critical to the validity of this approach for estimating target DNA copy number. In this review, we discuss the advances in development of digital PCR, the differences between digital PCR and real time quantitative PCR. We further show the advances in application of digital PCR in the clinical diagnosis, GMO analysis, single cell expression, environmental microbiology and next generation sequencing. The prospects for the applications of digital PCR are discussed.