title: Solid-support sample loading for DNA sequencing.
abstract: We present a new method for simplified low-quantity DNA loading onto microelectrophoresis devices. The method is based on combined solid-phase extraction, purification, and transport of DNA reversibly bound on paramagnetic microspheres. DNA is adsorbed onto the microspheres, captured with a magnetized permalloy wire, and then directly injected as a highly focused sample plug into the separation channel. This method circumvents both the minimum volume requirement of pipettors (since only solid beads are transported) and the timing complications of double-T microfluidic injection. Injections from Sanger samples of <100 pg total suspended weight match the signal strength of our previous conventional injections at >10-times the starting DNA sample. Sequencing traces show a resolution that matches or exceeds double-T injections. A kinetic model reproduces the time-dependence of the injection signals and proves that total nonidealities in the method produce injection-broadened plugs of approximately 1-s duration. The method should be broadly extendable to DNA and protein separations in both microdevice and capillary electrophoresis.
authors.affiliation: Whitehead Institute for Biomedical Research, Nine Cambridge Center, Cambridge, Massachusetts 02142, USA.
authors.forename: Jörn
authors.initials: J
authors.lastname: Ueberfeld
authors.forename: Sameh A
authors.initials: SA
authors.lastname: El-Difrawy
authors.forename: Korisha
authors.initials: K
authors.lastname: Ramdhanie
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Ehrlich
mesh_terms: D001483:Base Sequence; D004247:DNA / Q000032:analysis* / Q000378:metabolism; D004566:Electrodes; D004583:Electrons; D007700:Kinetics; D008855:Microscopy, Electron, Scanning; D008957:Models, Genetic
chemical_list: D004247:DNA