title: Universal CG cloning of polymerase chain reaction products.
abstract: Single-insert cloning of DNA fragments without restriction enzymes has traditionally been achieved using TA cloning, with annealing of a polymerase chain reaction (PCR) fragment containing a single overhanging 3' A to a plasmid vector containing a 3' T. In this article, we show that the analogous "CG cloning" is faster and far more efficient, using AhdI to generate a C-vector. For an afternoon ligation, CG cloning achieved double the cloning efficiency and more than 4-fold the number of transformants compared with TA cloning. However, blunt-end ligation was markedly more efficient than both. CG cloning could prove to be extremely useful for single-copy high-throughput cloning.
authors.affiliation: School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, NSW 2052, Australia; Department of Nutrition and Toxicology, University of California, Berkeley, CA 94720, USA.
authors.forename: Julian
authors.initials: J
authors.lastname: Stevenson
authors.affiliation: School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, NSW 2052, Australia. Electronic address: aj.brown@unsw.edu.au.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Brown
mesh_terms: D003001:Cloning, Molecular / Q000379:methods*; D004247:DNA / Q000235:genetics; D016133:Polymerase Chain Reaction; D013997:Time Factors
chemical_list: D004247:DNA
keywords: Cloning; PCR; Plasmid vectors