AQUA cloning: A versatile and simple enzyme-free cloning approach
11.09.2015
Beyer HM, Gonschorek P, Samodelov SL, Meier M, Weber W, Zurbriggen MD.
PLoS One. 2015;10(9):e0137652.
Assembly cloning is increasingly replacing conventional restriction enzyme and DNA- ligase-dependent cloning methods for reasons of efficiency and performance. Here, we describe AQUA (advanced quick assembly), a simple and versatile seamless assembly cloning approach. We demonstrate the applicability and versatility of AQUA Cloning in selected proof-of-principle applications including targeted insertion-, deletion- and site- directed point-mutagenesis, and combinatorial cloning. Furthermore, we show the one pot de novo assembly of multiple DNA fragments into a single circular plasmid encoding a com- plex light- and chemically-regulated Boolean A NIMPLY B logic operation. AQUA Cloning harnesses intrinsic in vivo processing of linear DNA fragments with short regions of homol- ogy of 16 to 32 bp mediated by Escherichia coli. It does not require any kits, enzymes or preparations of reagents and is the simplest assembly cloning protocol to date.
