SIMPLE Method for Plasmid Editing

Abstract

The development of molecular cloning techniques starting in the late 1960s created new tools to facilitate our understanding of gene function. Later advances in PCR and site-specific mutagenesis allowed researchers to efficiently dissect genes down to the single-nucleotide level. These methods have become virtually ubiquitous in nearly every biology laboratory. Recently, we developed an additional technique called SIMPLE (SapI/AarI incision-mediated plasmid editing), which allows for efficient deletions, short insertions (e.g., epitope tagging), and accurate mutations of episomal plasmids. SIMPLE cloning has a wide range of applications for molecular geneticists and adds a precision instrument to the biologist's toolbox. Here, we describe a detailed, step-by-step set of instructions for SIMPLE cloning.

Keywords: Molecular cloning; Plasmid editing; Site-directed mutagenesis.