Abstract
Gene targeting (GT) is an effective tool for functional genomics. We have developed an efficient and generally applicable homologous recombination (HR)-dependent GT procedure in rice, which comprises, (1) a large-scale Agrobacterium -mediated transformation, (2) a strong positive-negative selection, and (3) an efficient PCR screening to detect junction fragments generated by HR. To avoid any potential undesirable side effects, caused by manipulating recombination and/or repair system(s), for elucidating the function of a target gene, we employ wild-type rice cultivars for GT. Because independent GT events by HR should generate an identical genomic structure with a previously designed sequence alteration(s), the experimental demonstration of the capability for the reproducible isolation of recombinants with the anticipated gene structure would be very important. At present, we have reproducibly succeeded in generating rice GT plants of eight different genes (Waxy, Adh1, Adh2, OsDDM1a, OsDDM1bOsMET1a, OsMET1b, OsMET1a, OsMET1b, and OsDRM1a).
