New method to increase the efficiency of DNA extraction using dielectrophoresis

Abstract

Nucleic acid extraction is crucial for applications spanning from liquid biopsies to climate-resistant crop development. Dielectrophoresis has been described as possible alternative to the more laborious and toxic phenol-chloroform protocol currently employed, but so far, has been limited to analytical applications1, not being used as preparative technique due to poor yield, particularly for small fragments. The dielectrophoretic attraction is counteracted by the electrostatic and osmotic repulsion due to negative charges in the nucleic acids’ backbones. This prevents the high localized concentration as well as the aggregation required to capture smaller fragments. We tried to circumvent this challenge using principles from solid-phase reversible immobilization, namely: a) inducing DNA condensation using crowding agents, what will result in a configuration with less negative charges exposed and b) shielding the remaining charges using mono or di-valent cations2. We used a set of interdigitated gold electrodes, fabricate using conventional CMOS technology. Each electrode was 2 µm wide with a spacing of 2 µm. These dimensions were optimized to maximum capture of DNA. We started by investigating the role of monovalent cations by assessing the capture of lambda-DNA in buffer with increased concentration of Na+. The increased concentration of Na+ resulted in higher conductivity. However, at the frequency used (10 MHz), this has no impact in the real part of Clausius-Mossotti factor, hence on the dielectrophoretic force. The capture of lambda-DNA and retention after the electric field was turned off (measured by fluorescence) increased proportionally with the concentration of Na+ for an interval between 1.4 mM and 1400 mM. Next, we investigated the addition of IgG protein (100 mg/L) to the buffer with highest Na+ content. At physiological pH, IgG is positively charged and has a long molecular chain, able to induce crowding effect3. It presents only minimal negative dielectrophoretic response at the employed frequency. We observed that, 15 s after the field is applied, the capture of lambda-DNA reached a maximum in the control solution (not containing IgG), decreasing after that. In solutions containing the protein, the retention remained constant at the maximum value until the field was turned off. Additionally, DNA concentration after the field was turned off remained 2x higher in presence of IgG than in the control solution. Those initial experiments show a clear possibility of improving the performance of dielectrophoretic capture of DNA through manipulation of the sample composition.