MAGIC, an in vivo genetic method for the rapid construction of recombinant DNA molecules.
We describe a highly engineered in vivo cloning method, mating-assisted genetically integrated cloning (MAGIC), that facilitates the rapid construction of recombinant DNA molecules. MAGIC uses bacterial mating, in vivo site-specific endonuclease cleavage and homologous recombination to catalyze the transfer of a DNA fragment between a donor vector in one bacterial strain and a recipient plasmid in a separate bacterial strain. Recombination events are genetically selected and result in placement of the gene of interest under the control of new regulatory elements with high efficiency. MAGIC eliminates the need for restriction enzymes, DNA ligases, preparation of DNA and all in vitro manipulations required for subcloning and allows the rapid construction of multiple constructs with minimal effort. We show that MAGIC can generate constructs for expression in multiple organisms. As this new method requires only the simple mixing of bacterial strains, it represents a substantial advance in high-throughput recombinant DNA production that will save time, effort and expense in functional genomics studies.
Li
Mamie Z
MZ
Howard Hughes Medical Institute, Department of Genetics, Harvard Partners Center for Genetics and Genomics, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, Massachusetts 02115, USA.
Elledge
Stephen J
SJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods*; D004274:DNA, Recombinant / Q000235:genetics*; D018014:Gene Transfer Techniques; D008969:Molecular Sequence Data; D010957:Plasmids
D004274:DNA, Recombinant