Circular polymerase extension cloning of complex gene libraries and pathways.
High-throughput genomics and the emerging field of synthetic biology demand ever more convenient, economical, and efficient technologies to assemble and clone genes, gene libraries and synthetic pathways. Here, we describe the development of a novel and extremely simple cloning method, circular polymerase extension cloning (CPEC). This method uses a single polymerase to assemble and clone multiple inserts with any vector in a one-step reaction in vitro. No restriction digestion, ligation, or single-stranded homologous recombination is required. In this study, we elucidate the CPEC reaction mechanism and demonstrate its usage in demanding synthetic biology applications such as one-step assembly and cloning of complex combinatorial libraries and multi-component pathways.
Quan
Jiayuan
J
Department of Biomedical Engineering & Institute for Genome Sciences and Policy, Duke University, Durham, North Carolina, United States of America.
Tian
Jingdong
J
D003001:Cloning, Molecular; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D023281:Genomics; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic
D004259:DNA-Directed DNA Polymerase