Dna electrophoresis in photopolymerized polyacrylamide gels on a microfluidic device

• Main Author: Lo, Chih-Cheng
• Other Authors: Ugaz, Victor M.
• Format: Others
• Language: en_US
• Published: 2010
• Subjects: microchip electrophoresis; separation resolution; whole-column imaging detection
• Online Access: http://hdl.handle.net/1969.1/ETD-TAMU-2685; http://hdl.handle.net/1969.1/ETD-TAMU-2685

DNA gel electrophoresis is a critical analytical step in a wide spectrum of genomic analysis assays. Great efforts have been directed to the development of miniaturized microfluidic systems (“lab-on-a-chip” systems) to perform low-cost, high-throughput DNA gel electrophoresis. However, further progress toward dramatic improvements of separation performance over ultra-short distances requires a much more detailed understanding of the physics of DNA migration in the sieving gel matrix than is currently available in literature. The ultimate goal would be the ability to quantitatively determine the achievable level of separation performance by direct measurements of fundamental parameters (mobility, diffusion, and dispersion coefficients) associated with the gel matrix instead of the traditional trial-and-error process. We successfully established this predicting capability by measuring these fundamental parameters on a conventional slab gel DNA sequencer. However, it is difficult to carry out fast and extensive measurements of these parameters on a conventional gel electrophoresis system using single-point detection (2,000 hours on the slab gel DNA sequencer we used). To address this issue, we designed and built a new automated whole-gel scanning detection system for a systematic investigation of these governing parameters on a microfluidic gel electrophoresis device with integrated on-chip electrodes, heaters, and temperature sensors. With this system, we can observe the progress of DNA separation along the whole microchannel with high temporal and spatial accuracy in nearly real time. This is in contrast to both conventional slab gel imaging where the entire gel can be monitored, but only at one time frame after completion of the separation, and capillary electrophoresis systems that allows detection as a function of time, but only at a single detection location. With this system, a complete set of mobility, diffusion, and dispersion data can be collected within one hour instead of days or even months of work on a conventional sequencer under the same experimental conditions. The ability to acquire both spatial and temporal data simultaneously provides a more detailed picture of the separation process that can potentially be used to refine theoretical models and improve separation performance over ultra-short distances for the nextgeneration of electrophoresis technology.