Rapid one-step recombinational cloning.
As an increasing number of genes and open reading frames of unknown function are discovered, expression of the encoded proteins is critical toward establishing function. Accordingly, there is an increased need for highly efficient, high-fidelity methods for directional cloning. Among the available methods, site-specific recombination-based cloning techniques, which eliminate the use of restriction endonucleases and ligase, have been widely used for high-throughput (HTP) procedures. We have developed a recombination cloning method, which uses truncated recombination sites to clone PCR products directly into destination/expression vectors, thereby bypassing the requirement for first producing an entry clone. Cloning efficiencies in excess of 80% are obtained providing a highly efficient method for directional HTP cloning.
Fu
Changlin
C
Monsanto Company, 800 N. Lindbergh Blvd., St Louis, MO 63167, USA.
Wehr
Daniel R
DR
Edwards
Janice
J
Hauge
Brian
B
D001287:Attachment Sites, Microbiological; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods*; D017931:DNA Primers / Q000737:chemistry; D005822:Genetic Vectors; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D017931:DNA Primers