HOMOLOGOUS RECOMBINATION METHOD, CLONING METHOD, AND KIT

摘要

The present invention provides a method which can achieve the homologous recombination of a gene of interest selectively, and a recombinant DNA molecule produced by the method. A homologous recombination method which uses a PCR product and a linearlized vector is disposed. The PCR product comprises a sequence for a target gene and amplification primer sequences P1 and P2 on both terminal ends. The vector contains homologous recombination regions VP1 and VP2 which respectively comprises nucleotide sequences homologous to the amplification primer sequences P1 and P2 of the PCR product, and a homologous recombination region VT1 which comprises a nucleotide sequence homologous to a sequence T1 that is a part of sequence in the PCR product internal to P1 on the terminal side of VP1 and/or a homologous recombination region VT2 which comprises a nucleotide sequence homologous to a sequence T2 that is a part of sequence in the PCR product internal to P2 on the terminal side of VP2 (provided that at least one of T1 and T2 has a nucleotide sequence specific to the target gene). The method comprises subjecting the PCR product to a homologous recombination reaction to insert the PCR product into the vector, thereby producing a recombinant DNA molecule in which the PCR product of interest specifically inserted into the vector. Also disclosed a cloning method for producing the recombinant DNA molecule comprising the vector and the PCR product of interest specifically inserted into the vector by the homologous recombination method, and amplifying the recombinant DNA molecule.