Design and characterization of convective thermal cyclers for high-speed DNA analysis

Abstract

An ideal polymerase chain reaction (PCR) system should be capable of rapidly amplifying a wide range of targets in both single and multiplex formats. Unfortunately, the timescales and complexities involved in many existing technologies impose significant limitations on achievable throughput. Buoyancy driven PCR is emerging as a simplified version of thermally driven bio-analysis systems. Here, we demonstrate a simplified convectively driven thermocycler capable of performing single and multiplex PCR for amplicons ranging from 191 bp to 1.3 kb within 10 to 50 minutes using 10 to 25 µL reaction volumes. By positioning two independent thermoelectric heating elements along the perimeter of a flow loop reactor constructed using ordinary plastic tubing, a buoyancy-driven flow is established that continuously circulates reagents through temperature zones associated with the PCR process. Unlike conventional benchtop thermocyclers, this arrangement allows reactions to be performed without the need for dynamic temperature control of inactive hardware components while maintaining comparable product yields and requiring no modifications to standard PCR protocols. We also provide a general correlation that can be applied to design reactor geometries satisfying virtually any combination of reagent volume and cycling time. In addition to offering an attractive combination of cost and performance, this system is readily adaptable for portable battery powered operation, making it feasible to perform PCRbased assays in a broader array of settings.