(185a) Dielectrophoretic Sorting of Plasmid and Genomic DNA

Innovations in DNA sample preparation may soon enable development of rapid sequencing and bioanalysis tools that significantly improve the availability and accuracy of nucleic acid analysis. Over the years, a variety of methods have been used to separate nucleic acid fragments based on size. Since various Next-Generation Sequencing (NGS) methods require DNA fragments of specific size, the need for improved sorting has fueled significant analytical research. Achieving efficient and continuous flow fractionation of DNA and RNA still presents considerable challenges. Current separation methods have not fully satisfied the need for rapid fractionation of heterogeneous nucleic acid samples into populations of specific and narrow size distribution. We have recently explored the use of dielectrophoresis to improve sized based, continuous sorting with reduced analysis times. For this purpose, we have developed a microfluidic constriction-sorter device, using insulator-based dielectrophoresis and pressure-driven flow. This approach enables continuous-flow separation of nucleic acid analytes into physically distinct microchannel outlets. We demonstrate AC dielectrophoretic sorting of four different DNA analytes, including both plasmid and genomic DNA. The sorting behavior is tunable and size-specific. It demonstrates strong dependency on applied potentials and frequencies. Specifically, we have investigated the behavior of 1.0, 10.2, 19.5, and 49.5 kbp dsDNA. Applied potentials ranged from 200 to 2400 V/cm, while frequencies ranged from 50 Hz to 20 kHz with flow rates in the range of 1 µL/h. These conditions yielded a maximum observed sorting efficiency of approximately 92%. Based on these results, we propose the use of a continuous-flow fractionation strategy to facilitate future coupling with NGS approaches.