Nature Biotechnology
14, 1123 - 1128 (1996)
doi:10.1038/nbt0996-1123
A strategy for rapid and efficient DNA sequencing by mass spectrometryHubert Köster1, *, Kai Tang2, Dong-Jing Fu2, Andreas Braun3, Dirk van den Boom1, Cassandra L. Smith5, Robert J. Cotter4
& Charles R. Cantor5
1University of Hamburg, Department of Biochemistry and Molecular Biology, Martin-Luther-King-Platz 6, 20146 Hamburg, Germany.
2Sequenom Inc., 101 Arch St., Boston, MA 02110, USA.
3Sequenom Instruments GmbH, Mendelssohnstr. 15D, 22761 Hamburg, Germany.
4Johns Hopkins University, School of Medicine, Department of Pharmacology, 725 N. Wolfe St., Baltimore, MD 21205, USA.
5Boston University, Center for Advanced Biotechnology, 36 Cummington St., Boston, MA 02215, USA.
*e-mail: koester@chemie.uni-hamburg.d400.de Two methods of solid-phase Sanger DNA sequencing followed by detection with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry are demonstrated. In one method, sequencing ladders generated on an immobilized synthetic template were resolved up to the 63-mer including the primer. Detection sensivity and resolution were sufficient for sequence analysis in the given range. This approach is particularly suitable for comparative (diagnostic) DNA sequencing. A second method that has the potential for high throughput de novo DNA sequencing is also presented; it uses immobilized duplex probes with five-base single-stranded overhangs to capture an unknown DNA template serving as primers for Sanger DNA sequencing. The power of mass spectrometry is demonstrated not only by its very high speed, but also by its ability to identify sequences that are not readable using gel electrophoresis.
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