A highly parallel method for synthesizing DNA repeats enables the discovery of 'smart' protein polymers.

Published

Journal Article

Robust high-throughput synthesis methods are needed to expand the repertoire of repetitive protein-polymers for different applications. To address this need, we developed a new method, overlap extension rolling circle amplification (OERCA), for the highly parallel synthesis of genes encoding repetitive protein-polymers. OERCA involves a single PCR-type reaction for the rolling circle amplification of a circular DNA template and simultaneous overlap extension by thermal cycling. We characterized the variables that control OERCA and demonstrated its superiority over existing methods, its robustness, high-throughput and versatility by synthesizing variants of elastin-like polypeptides (ELPs) and protease-responsive polymers of glucagon-like peptide-1 analogues. Despite the GC-rich, highly repetitive sequences of ELPs, we synthesized remarkably large genes without recursive ligation. OERCA also enabled us to discover 'smart' biopolymers that exhibit fully reversible thermally responsive behaviour. This powerful strategy generates libraries of repetitive genes over a wide and tunable range of molecular weights in a 'one-pot' parallel format.

Full Text

Duke Authors

Cited Authors

  • Amiram, M; Quiroz, FG; Callahan, DJ; Chilkoti, A

Published Date

  • February 2011

Published In

Volume / Issue

  • 10 / 2

Start / End Page

  • 141 - 148

PubMed ID

  • 21258353

Pubmed Central ID

  • 21258353

International Standard Serial Number (ISSN)

  • 1476-1122

Digital Object Identifier (DOI)

  • 10.1038/nmat2942

Language

  • eng

Conference Location

  • England