Recombinational cloning using nucleic acids having recombination sites and methods for synthesizing double stranded nucleic acids

摘要

A method for synthesising a double stranded nucleic acid molecule comprising: a) mixing one or more nucleic acid templates with a polypeptide having polymerase activity and one ore more primers comprising at least a first recombination site or functional portions thereof; b) incubating said mixture under conditions sufficient to synthesise a first nucleic acid molecule which is complementary to all or a functional portion of said one or more templates and which comprises at least said first recombination site or functional portions thereof; and c) incubating said first nucleic acid molecule in the presence of one or more primers comprising at least a second recombination site or functional portions thereof under conditions sufficient to synthesise a second nucleic acid molecule complementary to all or a functional portion to said first nucleic acid molecule, thereby producing a double stranded nucleic acid molecule comprising at least said first and second recombination sites or functional portions thereof; wherein at least one of said first and second recombination sites comprises at least one nucleotide sequence that has at least 80-99% homology to a nucleotide sequence selected from a group of sequences disclosed.