Direct droplet digital PCR (dddPCR) for species specific, accurate and precise quantification of bacteria in mixed samples

Abstract

Here we demonstrate a direct droplet digital PCR assay (dddPCR) for simultaneous identification and quantification of bacteria in a bacterial mixture. We encapsulate bacterial cells in droplets together with PCR reagents to amplify specific target genes. Our method introduces accurate and precise quantitation of bacteria in a bacterial mixture by digital PCR, while eliminating the errors and workload associated with extraction and purification of DNA. The method thus combines the precision and specificity of digital PCR with robustness of a direct PCR. We also show that dddPCR is more tolerant to inhibition by non-specific bacteria as compared to traditional direct real-time PCR.