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Originally published in Science Express on 4 August 2005
Science 9 September 2005:
Vol. 309. no. 5741, pp. 1728 - 1732
DOI: 10.1126/science.1117389
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Reports

Accurate Multiplex Polony Sequencing of an Evolved Bacterial Genome

Jay Shendure,1*{dagger} Gregory J. Porreca,1*{dagger} Nikos B. Reppas,1 Xiaoxia Lin,1 John P. McCutcheon,2,3 Abraham M. Rosenbaum,1 Michael D. Wang,1 Kun Zhang,1 Robi D. Mitra,2 George M. Church1

We describe a DNA sequencing technology in which a commonly available, inexpensive epifluorescence microscope is converted to rapid nonelectrophoretic DNA sequencing automation. We apply this technology to resequence an evolved strain of Escherichia coli at less than one error per million consensus bases. A cell-free, mate-paired library provided single DNA molecules that were amplified in parallel to 1-micrometer beads by emulsion polymerase chain reaction. Millions of beads were immobilized in a polyacrylamide gel and subjected to automated cycles of sequencing by ligation and four-color imaging. Cost per base was roughly one-ninth as much as that of conventional sequencing. Our protocols were implemented with off-the-shelf instrumentation and reagents.

1 Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
2 Department of Genetics, Washington University, St. Louis, MO 63110, USA.
3 Howard Hughes Medical Institute, Washington University, St. Louis, MO 63110, USA.

* These authors contributed equally to this work.

{dagger} To whom correspondence should be addressed. E-mail: shendure{at}alumni.princeton.edu (J.S.), gregory_porreca{at}student.hms.harvard.edu (G.J.P.)

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