Scalable amplification of strand subsets from chip-synthesized oligonucleotide libraries

Thorsten L. Schmidt, Brian J. Beliveau, Yavuz O. Uca, Mark Theilmann, Felipe Da Cruz, Chao-Ting Wu and William M. Shih ()
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Thorsten L. Schmidt: Dana-Farber Cancer Institute
Brian J. Beliveau: Harvard Medical School
Yavuz O. Uca: Center for Advancing Electronics Dresden (cfaed), Technische Universität Dresden
Mark Theilmann: Dana-Farber Cancer Institute
Felipe Da Cruz: Wyss Institute for Biologically Inspired Engineering at Harvard
Chao-Ting Wu: Harvard Medical School
William M. Shih: Dana-Farber Cancer Institute

Nature Communications, 2015, vol. 6, issue 1, 1-7

Abstract: Abstract Synthetic oligonucleotides are the main cost factor for studies in DNA nanotechnology, genetics and synthetic biology, which all require thousands of these at high quality. Inexpensive chip-synthesized oligonucleotide libraries can contain hundreds of thousands of distinct sequences, however only at sub-femtomole quantities per strand. Here we present a selective oligonucleotide amplification method, based on three rounds of rolling-circle amplification, that produces nanomole amounts of single-stranded oligonucleotides per millilitre reaction. In a multistep one-pot procedure, subsets of hundreds or thousands of single-stranded DNAs with different lengths can selectively be amplified and purified together. These oligonucleotides are used to fold several DNA nanostructures and as primary fluorescence in situ hybridization probes. The amplification cost is lower than other reported methods (typically around US$ 20 per nanomole total oligonucleotides produced) and is dominated by the use of commercial enzymes.

Date: 2015
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DOI: 10.1038/ncomms9634

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