A high-throughput analytical method for multiple DNA targets based on microdroplet PCR coupled with DGGE

Abstract

We developed a novel approach to analyze multiple DNA targets based on microdroplet PCR coupled with denaturing gradient gel electrophoresis (MPDG) to achieve high-throughput detection of biological samples. The target DNAs were preamplified using specific primers. Subsequently, the preamplified products were separated into individual microreactors for parallel amplification with high efficiency, avoiding the interference of different primers and templates, and preventing inconsistent amplification efficiency and non-specific amplification. The final products were analyzed using denaturing gradient gel electrophoresis (DGGE). Using genetically modified (GM) maize as samples, the MPDG method could be used for the simultaneous detection of three DNA targets with an absolute limit of detection of 0.5% (w/w), with no cross reaction with other non-GM samples. The simulated sample assay of MPDG suggested that the method is suitable for practical application. The MPDG approach, with high sensitivity and specificity, could play a crucial role in the field of nucleic acid detection.