Electrostatic Purification of Nucleic Acids for Micro Total Analysis Systems

摘要

Nucleic acids (such as DNA and RNA) play a central role in biological systems, carrying instructions that govern the function of all living organisms. Many of the most promising applications of lab-on-a-chip technology, such as infectious disease detection, gene expression profiling and DNA sequencing and amplification involve processing these carriers of genetic information. While decades of work has been dedicated to moving processes like these on-chip, the important precursor steps of isolating nucleic acids from samples have been largely overlooked. Fully integrating the sample preparation on-chip will increase processing speed while reducing operator error and sample contamination as well as labor and reagent related costs. To date the typical approach for implementing nucleic acid purification in a microchannel format is to translate the commonly used solid phase extraction method which relies on reagent changes to achieve immobilization and release of DNA. By using electrostatic control of DNA we are able to avoid the reagents (including PCR inhibitors) and complicated valving that this approach requires. We microfabricate chips using photolithography to create gold microelectrodes on glass substrates, upon which we bond a PDMS microchannel which is fabricated using soft lithography. Through COMSOL simulations we optimize our electrode geometry to maximize DNA capture efficiency. Designs are qualitatively evaluated using fluorescence microscopy. For quantification, we measure both the DNA removed from a sample (capture efficiency) and the DNA that can be released back into solution (elution efficiency) using UV spectrophotometry. By dramatically improving the ease and flexibility of nucleic acid purification, this enhanced microfluidic module can be integrated into existing microfluidic designs, improving how researchers approach on-chip nucleic acid processing and diagnostics.