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Molecular Pathology

DNA sequencing

Real-time single-molecule DNA sequence determination


Author: Rodney E. Shackelford, D.O., Ph.D., Tulane University (see Reviewers page)
Revised: 13 February 2011, last major update February 2011
Copyright: (c) 2008-2011, PathologyOutlines.com, Inc.

General
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● VisiGen Biotechnologies has engineered a DNA polymerase which acts as a real-time sensor of modified nucleotides
● This method uses DNA synthesis, employing a DNA polymerase with a donor fluorescent dye close to the polymerase active site where nucleotides to be incorporated are selected
● The four dNTPs are also modified, with each having a different acceptor dye compatible with the polymerase donor dye
● At dNTP incorporation, the donor dye transfers fluorescent resonant energy to the acceptor dye, resulting in measurable light release
● The four different acceptor dyes give off different wave length emissions, allowing the specific dNTP incorporated to be determined
● The advantage of this system is that it can sequence DNA at the same rate at the polymerase incorporates dNTPs - several hundred moieties per second, with read-lengths of around 1 kb, longer than possible with any current platform
● This technology may eventually allow incredible sequencing feats, such a sequencing the entire human genome in one day for ~$1,000 (N Biotechnol 2009;25:195)

End of Molecular Pathology > DNA sequencing > Real-time single-molecule DNA sequence determination


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