Application of droplet digital PCR to detect the pathogens of infectious diseases

Abstract

Polymerase chain reaction (PCR) is a molecular biology technique used to multiply certain deoxyribonucleic acid (DNA) fragments. It is a common and indispensable technique that has been applied in many areas, especially in clinical laboratories. The third generation of polymerase chain reaction, droplet digital polymerase chain reaction (ddPCR), is a biotechnological refinement of conventional polymerase chain reaction methods that can be used to directly quantify and clonally amplify DNA. Droplet digital polymerase chain reaction is now widely used in low-abundance nucleic acid detection and is useful in diagnosis of infectious diseases. Here, we summarized the potential advantages of droplet digital polymerase chain reaction in clinical diagnosis of infectious diseases, including viral diseases, bacterial diseases and parasite infections, concluded that ddPCR provides a more sensitive, accurate, and reproducible detection of low-abundance pathogens and may be a better choice than quantitative polymerase chain reaction for clinical applications in the future.

Keywords: Droplet digital PCR; Polymerase chain reaction; diagnosis; infectious disease.

Figure 1. Comparisons of the flowchart and principle of ddPCR and qPCR

In droplet digital PCR, the sample is partitioned into 20,000 nanoliter-sized droplets that were individually analyzed. But for qPCR, a single sample offers only a single measurement like traditional PCR. This partitioning in ddPCR process enables the measurement of thousands of independent amplification events within a single sample, so the droplet digital PCR provides an absolute count of target DNA copies per sample without the standard curve when qPCR calculates the target DNA number based on standard curve.