An efficient and accurate integration of mini-Mu transposons in vitro: a general methodology for functional genetic analysis and molecular biology applications

Transposons are mobile genetic elements and have been utilized as essential tools in genetics over the years, Though highly useful, many of the curren t transposon-based applications suffer from various limitations, the most n otable of which are: (i) transposition is performed in vivo, typically spec ies specifically, and as a multistep process; (ii) accuracy and/or efficien cy of the in vivo or in vitro transposition reaction is not optimal; (iii) a limited set of target sites is used. We describe here a genetic analysis methodology that is based on bacteriophage Mu DNA transposition and circumv ents such limitations. The Mu transposon tool is composed of only a few com ponents and utilizes a highly efficient and accurate in vitro DNA transposi tion reaction with a low stringency of target preference, The utility of th e Mu system in functional genetic analysis is demonstrated using restrictio n analysis and genetic footprinting strategies. The Mu methodology is readi ly applicable in a variety of current and emerging transposon-based techniq ues and is expected to generate novel approaches to functional analysis of genes, genomes and proteins.