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Journal of Nucleic Acids
Volume 2012, Article ID 923214, 11 pages
Review Article

Directed Evolution of Proteins through In Vitro Protein Synthesis in Liposomes

1ERATO Japan Science and Technology (JST) and Yomo Dynamical Micro-Scale Reaction Environment Project, Graduate School of Information Sciences and Technology, Osaka University, 1-5 Yamadaoka, Suita, Osaka 565-0871, Japan
2Graduate School of Information Science and Technology, Osaka University, 1-5 Yamadaoka, Suita, Osaka 565-0871, Japan
3Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan
4Graduate School of Frontier Biosciences, Osaka University, 1-5 Yamadaoka, Suita, Osaka 565-0871, Japan

Received 8 June 2012; Accepted 10 July 2012

Academic Editor: Hiroshi Murakami

Copyright © 2012 Takehiro Nishikawa et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


Directed evolution of proteins is a technique used to modify protein functions through “Darwinian selection.” In vitro compartmentalization (IVC) is an in vitro gene screening system for directed evolution of proteins. IVC establishes the link between genetic information (genotype) and the protein translated from the information (phenotype), which is essential for all directed evolution methods, by encapsulating both in a nonliving microcompartment. Herein, we introduce a new liposome-based IVC system consisting of a liposome, the protein synthesis using recombinant elements (PURE) system and a fluorescence-activated cell sorter (FACS) used as a microcompartment, in vitro protein synthesis system, and high-throughput screen, respectively. Liposome-based IVC is characterized by in vitro protein synthesis from a single copy of a gene in a cell-sized unilamellar liposome and quantitative functional evaluation of the synthesized proteins. Examples of liposome-based IVC for screening proteins such as GFP and β-glucuronidase are described. We discuss the future directions for this method and its applications.