Construction of an Ultra-Large Phage Display Library by Kunkel Mutagenesis and Rolling Circle Amplification.
Methods in molecular biology (Clifton, N.J.)(2023)
摘要
An important contributor to the successful generation of recombinant affinity reagents via phage display is a large and diverse library. We describe, herein, the application of Kunkel mutagenesis and rolling circle amplification (RCA) to the construction of a 1.1 × 10 member library, with only 26 electroporations, and isolation of low- to sub-nanomolar monobodies to a number of protein targets, including human COP9 signalosome subunit 5 (COPS5), HIV-1 Rev. binding protein-like protein (HRBL), X-ray repair cross-complementing 5/6 (Ku70/80) heterodimer, the receptor-binding domain (RBD) of SARS-CoV-2, and transforming growth factor beta 1 (TGF-β1).
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关键词
kunkel mutagenesis,rolling circle amplification,ultra-large
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