Accessing DNA by low voltage alternating current Joule effect heating

A DNA release sample preparation method based on the use! of low voltage alternating currents (LVACs) to generate Joule effect heating (JEH) is reported. This is a simple cell disruption strategy that offers internal, homogenous, rapid and low power consumption heating for the access of analytical grade DNA in seconds. A 100 muL JEH microreactor with a parallel and symmetric two electrode arrangement for uniform field generation was fabricated by machining and used to characterise JEH and DNA release from human epithelia, yeast (Saccharomyces cerevisiae) and Gram-positive bacteria (Enterococcus faecium) cell types. A 1 kHz sinusoidal low voltage (e.g. 10 Vrms) alternating current was used to reduce electrode: sample interactions. Following 96degreesC JEH treatment, effective DNA release was identified by PicoGreen(R) quantification for all three cell types. The JEH treated sample material was further successfully used, without purification, as a PCR template. Exposure to JEH-mediated 96degreesC temperatures for a 1 s duration was used to provide PCR-grade DNA template material from S. cerevisiae and E. faecium cells, and a 10 s duration was used for human epithelia cells. However, prolonged (> 1 min) exposure to 96degreesC JEH-mediated temperatures resulted in diminished DNA returns and the production of components that interfered with the PCR reaction. Further miniaturisation of the LVAC JEH cell by microfabtication was considered, and a JEH microreactor designs were evaluated by FLOTHERM v3.2 thermal modelling. Thermal isolation, using a free-standing cavity structure was identified as an excellent means to enable rapid heating (220degreesC s(-1)) with low power consumption (0.2 W). (C) 2004 Elsevier B.V. All rights reserved.