UNIT 3.20 Site-Specific Recombinational Cloning Using Gateway and In-Fusion Cloning Schemes

The comprehensive study of protein structure and function, or proteomics, depends on the obtainability of full-length cDNAs in species-specific expression vectors and subsequent functional analysis of the expressed protein. Recombinational cloning is a universal cloning technique based on site-specific recombination that is independent of the insert DNA sequence of interest, which differentiates it from classical restriction enzyme−based cloning methods. Recombinational cloning enables rapid and efficient parallel transfer of DNA inserts into multiple expression systems. This unit summarizes strategies for generating expression-ready clones using the most popular commercial recombinational cloning technologies, Gateway (Life Technologies) and In-Fusion (Clontech). © 2015 by John Wiley & Sons, Inc. Keywords: recombinational cloning; Gateway; In-Fusion; high-throughput; Cre-Lox