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Updated: Jun 1, 2026

Mutagenesis and Functional Selection Protocols for Directed Evolution of Proteins in E. coli
Published on: March 16, 2011
Beyond natural evolution: multi-scale in vivo mutagenesis toolkits for synthetic evolution
Zhijun Teng1, Julian E Prieto-Vivas2, Kevin J Verstrepen3
1State Key Laboratory of Engineering Biology for Low-Carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China; Haihe Laboratory of Synthetic Biology, Tianjin 300308, China; University of Chinese Academy of Sciences, Beijing 100049, China; National Center of Technology Innovation for Synthetic Biology, Tianjin 300308, China; TIB - VIB Joint Center of Synthetic Biology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
Abstract:
Our current industrial, agricultural, and medical practices exploit the extraordinary biodiversity generated through billions of years of natural evolution. Despite their high fitness in native habitats, biomolecules and organisms are often not optimally suited for industrial and medical use. Synthetic evolution leverages technologies such as DNA synthesis, CRISPR-Cas engineering, and synthetic biology to enable continuous in vivo mutagenesis of biomolecules or organisms to improve specific desirable characteristics. This review presents the latest mutagenesis toolkits classified by mutational scale: genome-wide, medium-scale, and site-specific, each tailored to different application scenarios. We discuss the mechanisms and capabilities underlying each scale, analyze current limitations, and highlight the untapped potential of next-generation gene-editing technologies, high-throughput screening, and artificial intelligence in advancing synthetic evolution.
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