To develop the gene targeting system by homologous recombination in Arabidopsis thaliana, we constructed two targeting vectors and showed the reliability of the scheme which is based on genetic complementation of phosphoribosylanthranilate transferase (PAT1) gene. The PAT1 gene, which is essential for tryptophan biosynthesis, was selected as a target gene because the loss of funciton leads to fluorescence pheonotype due to the accumulation of anthranilic acid derivatives. pHS113 contains PAT1 gene is replaced by the neomycin phosphotransferase gene (NPTII). pHS117 consists of 1.1 kbinternal fragment of genomic PAT1 gene following with NPTII gene. In this targeting strategy, Arabidopsis PAT1 gene can be disrupted by single-step of transformation experiment.