A strategy for rapid and efficient DNA sequencing by mass spectrometry

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 pri...

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Bibliographic Details
Published in:Nature biotechnology 1996-09, Vol.14 (9), p.1123-1128
Main Authors: Köster, Hubert, Tang, Kai, Fu, Dong-Jing, Braun, Andreas, van den Boom, Dirk, Smith, Cassandra L, Cotter, Robert J, Cantor, Charles R
Format: Article
Language:English
Subjects:
Agriculture
Base Sequence
Bioinformatics
Biological and medical sciences
Biomedical and Life Sciences
Biomedical Engineering/Biotechnology
Biomedicine
Biotechnology
DNA - genetics
DNA Primers - genetics
Evaluation Studies as Topic
Fundamental and applied biological sciences. Psychology
Genetic engineering
Genetic technics
Life Sciences
Methods. Procedures. Technologies
Molecular Sequence Data
research-article
Sensitivity and Specificity
Sequence Analysis, DNA - methods
Sequence Analysis, DNA - statistics & numerical data
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization - methods
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization - statistics & numerical data
Synthetic digonucleotides and genes. Sequencing
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Summary: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.
ISSN:1087-0156
1546-1696
DOI:10.1038/nbt0996-1123