Microdevice-based measurements of diffusion and dispersion in cross-linked and linear polyacrylamide DNA sequencing gels.
We use microfabricated gel electrophoresis devices incorporating integrated on-chip electrodes, heaters, and temperature sensors to measure diffusion and dispersion of single-stranded DNA fragments in cross-linked and uncross-linked polyacrylamide gels. The microdevice format allows a complete set of diffusion and dispersion data to be collected in approximately one hour. These results are compared with corresponding data obtained in a macroscale DNA sequencer, and the effects of gel composition and initiation chemistry are explored. Although the diffusion and dispersion data exhibit similar qualitative trends both on chip and on the macroscale, the magnitudes of the coefficients measured in the microdevice are somewhat higher. This discrepancy is likely due to altered polymerization kinetics arising as a consequence of using a UV-initiated polymerization chemistry to cast the on-chip gels as opposed to the standard chemical polymerization employed on the macroscale. We also find that reductions in the magnitudes of diffusion and dispersion coefficients are achieved at higher polymer concentrations and at operating temperatures in the vicinity of 50 degrees C. Finally, we find that cross-linked polyacrylamide gels yield significantly lower diffusion and dispersion coefficients than linear polyacrylamide. These findings can be used to identify rational strategies to improve separation performance in both micro- and macroscale gel electrophoresis systems.
Ugaz
Victor M
VM
Department of Chemical Engineering, The University of Michigan, Ann Arbor, MI 48109-2136, USA.
Burke
David T
DT
Burns
Mark A
MA
D003432:Cross-Linking Reagents; D004058:Diffusion; D004591:Electrophoresis, Polyacrylamide Gel / Q000295:instrumentation*; D004867:Equipment Design; D008832:Microchemistry / Q000295:instrumentation*; D008904:Miniaturization; D010777:Photochemistry; D017422:Sequence Analysis, DNA / Q000295:instrumentation*; D012825:Silicon; D013696:Temperature
D003432:Cross-Linking Reagents; D012825:Silicon