DNA virus extraction using the vortex fluidic device (VFD)

摘要

Currently, DNA extraction methods have a number of drawbacks. These are mainly related to the use of hazardous chemicals in the liquid-based method, high costs and the non-reusable spin column kits for the solid-phase method. Here, research has been conducted to attempt to extract DNA out of bacteriophage ??X174 as a model virus. In this method, high shear stress was imparted onto the phage through the use of a vortex fluidic device (VFD). The aims of the study were to simplify the process of viral DNA extraction. We attempted to use the high shear forces in the VFD to shear open the capsid. This process would result in the release of DNA into solution and an empty, intact capsid after the process. For this work, bacteriophage ??X174 was processed under a wide range of VFD processing parameters. These include the use of variable sample tube sizes at different sample tube tilt angles: (0??, 45??, and 90??). Importantly, the rotational speed of the sample tube was varied (2000-8000 rpm) and studies into the effect of vibrations on the fluid flow. In a novel approach, the bacteriophage was exposed to laser radiation combined with high levels of shear force. The effect of VFD-mediated processing was assessed by six techniques including: UV-Vis spectroscopy (in-situ and post), dynamic light scattering, atomic force microscopy, scanning electron microscopy, flow cytometry and gel electrophoresis. The results obtained suggest that there is phage fragmentation occurred during the VFD-mediated process, but DNA in solution was not being detected. This implied that the DNA in solution might be in a too lower concentration for the detection method we employed. Therefore, further studying this VFD-mediated process for viral DNA extraction is still feasible and should be continued to create an analysis that is suited for low concentrations of DNA.