| Autor: |
Bryan Crane, Rudolf Gilmanshin, Nanor Kojanian, Nicaulas Sabourin, Richard Allen, Yi Zhou, Jeffrey R. Krogmeier, Qun Zhong, Kedar Vyavahare, Saad Shaikh, Kate Carson, Jonathan W. Larson, Linda Knaian |
| Rok vydání: |
2009 |
| Předmět: |
|
| Zdroj: |
Biophysical Journal. 96(3) |
| ISSN: |
0006-3495 |
| DOI: |
10.1016/j.bpj.2008.12.1576 |
| Popis: |
Direct Linear Analysis (DLA) technology obtains high content sequence information by optically mapping sequence specific fluorescent tags bound to elongated genomic DNA molecules in shear flows.[1] To facilitate sensitivity and throughput, we have implemented a high molecular weight DNA concentrating system by photopatterning a semi-permeable membrane inside the microfluidic device. This minimizes the fluid volume in which the DNA molecules reside prior to optical mapping leading to decreased read times. The membrane is selectively permeable to buffer ions but not high molecular weight DNA molecules allowing enhanced sample concentration at the membrane surface during electrokinetic transfer. In addition, the semi-permeable membrane allows electrophoretic sample transfer into and throughout the microfluidic device avoiding hydrodynamic induced shear forces that can degrade the integrity of large DNA molecules. The device employs novel microfluidic channel geometries to limit the electric field strength to appropriate levels near the membrane surface to minimize both sample and membrane degradation while maintaining a sufficiently high electric field for rapid sample transfer. Additionally, ion polarization across the membrane due to selective membrane permeability is addressed by active buffer replenishment through devoted channels behind the membrane. This architecture is amenable to integration into electrophoretic systems requiring rapid sample concentration, positioning, and transfer between microfluidic components. This research was supported by the Department of Homeland Security Science and Directorate Technology.[1] Chan et al., Genome Research, 2004, 14:1137. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
|
