DOE Human Genome Program Contractor-Grantee
37. DNA Sequencing via Electrospray and Ion/Ion Chemistry in an Electrodynamic Ion Trap
Scott A. McLuckey1, James L. Stephenson, Jr.2, and Gregory B. Hurst2
1Department of Chemistry, Purdue University, West Lafayette, IN 47907-1393 and 2Chemical and Analytical Sciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831
We are pursuing a methodology for high speed DNA sequencing based on electrospray ionization mass spectrometry employing gas-phase ion/ion chemistry in a quadrupole ion trap. DNA sequencing via mass spectrometry has been pursued by a number of groups in recent years due to its promise for the obviation of time-consuming electrophoresis-based separations required with established sequencing strategies. By far, most effort has been directed toward matrix-assisted laser desorption ionization (MALDI) combined with time-of-flight mass spectrometry. While a MALDI-based approach may yet fulfill its promise, limitations encountered in ionizing relatively large DNA oligomers have proved to be difficult to overcome. In contrast, ionization of large DNA oligomers is not a limitation for electrospray ionization. However, electrospray-based approaches for high speed DNA sequencing have not been extensively pursued due to spectral congestion associated with the multiple charging phenomenon that is characteristic of electrospray. The formation of multiple charge states from a single oligomer severely limits the mixture complexity amenable to direct analysis via electrospray. For this reason, electrospray usually follows a separation method, such as liquid chromatography or capillary electrophoresis, when applied to mixtures. We have recently shown that gas-phase ion/ion chemistry involving oppositely charged ions within a quadrupole ion trap greatly expands the mixture analysis capability of electrospray. In this work, the idea is to subject Sanger mixtures to electrospray and ion/ion chemistry as a core element in a strategy for high speed DNA sequencing. This talk describes the methodology and progress to date.
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