A method was developed for efficiently transforming Arabidopsis thaliana root explants using Agrobacterium tumefaciens and kanamycin selection. Root explants were cultured to regenerate shoots rapidly and at 100% efficiency. Transformed seed-producing plants were obtained with an efficiency between 20% and 80% within 3 months after gene transfer. F1 seedlings showed Mendelian segregation of the kanamycin resistance trait. The method was applicable to three different Arabidopsis ecotypes. A chimeric bar gene conferring resistance to the herbicide Basta was introduced into Arabidopsis, and its expression was confirmed by enzymatic assay. Arabidopsis thaliana is a valuable model organism for plant molecular biology due to its small genome size, rapid generation time, and excellent genetics. However, the lack of a rapid and efficient transformation procedure was a major obstacle in gene-cloning strategies. Several transformation procedures using leaf material infected by Agrobacterium tumefaciens have been published, but regeneration was generally inefficient and took 4–5 months or more. Therefore, the regenerative response of Arabidopsis roots was examined. It was found that Arabidopsis root explants have a high potential for rapid shoot regeneration. Fertile plants can be regenerated reproducibly within 2–3 months. This regeneration method was combined with Agrobacterium infection to develop an efficient and rapid transformation procedure using kanamycin selection. The transformation procedure involved the use of a nononcogenic tumor-inducing Ti plasmid carrying a chimeric neomycin phosphotransferase II (neo) gene and a chimeric bar gene. The neo gene encodes neomycin phosphotransferase II (NeoPTase II), and the bar gene confers resistance to the herbicide Basta. The transformation method was tested with three different Arabidopsis ecotypes. DNA from four randomly chosen KmR regenerants was analyzed by DNA gel-blot hybridization to determine the T-DNA copy number. The data suggested that each of three transformants had one T-DNA insert hybridizing to both right- and left-border probes. However, one transformant probably contained four inserts. One DNA fragment of this transformant hybridized to both right- and left-border probes, suggesting that two of the four inserts might be arranged in a directly repeated tandem structure. In addition, both NeoPTase II and PAcTase activities were found in all transformants tested. The neo gene segregation in the F1 progeny of transformants confirmed the low-copy-number T-DNA insertion. Seeds from 10 randomly chosen transformants were germinated on GM containing 50 mg of Km per liter. Two weeks after germination, KmS seedlings had completely etiolated cotyledons, and development of roots and leaves was completely inhibited. By contrast, KmR plantsA method was developed for efficiently transforming Arabidopsis thaliana root explants using Agrobacterium tumefaciens and kanamycin selection. Root explants were cultured to regenerate shoots rapidly and at 100% efficiency. Transformed seed-producing plants were obtained with an efficiency between 20% and 80% within 3 months after gene transfer. F1 seedlings showed Mendelian segregation of the kanamycin resistance trait. The method was applicable to three different Arabidopsis ecotypes. A chimeric bar gene conferring resistance to the herbicide Basta was introduced into Arabidopsis, and its expression was confirmed by enzymatic assay. Arabidopsis thaliana is a valuable model organism for plant molecular biology due to its small genome size, rapid generation time, and excellent genetics. However, the lack of a rapid and efficient transformation procedure was a major obstacle in gene-cloning strategies. Several transformation procedures using leaf material infected by Agrobacterium tumefaciens have been published, but regeneration was generally inefficient and took 4–5 months or more. Therefore, the regenerative response of Arabidopsis roots was examined. It was found that Arabidopsis root explants have a high potential for rapid shoot regeneration. Fertile plants can be regenerated reproducibly within 2–3 months. This regeneration method was combined with Agrobacterium infection to develop an efficient and rapid transformation procedure using kanamycin selection. The transformation procedure involved the use of a nononcogenic tumor-inducing Ti plasmid carrying a chimeric neomycin phosphotransferase II (neo) gene and a chimeric bar gene. The neo gene encodes neomycin phosphotransferase II (NeoPTase II), and the bar gene confers resistance to the herbicide Basta. The transformation method was tested with three different Arabidopsis ecotypes. DNA from four randomly chosen KmR regenerants was analyzed by DNA gel-blot hybridization to determine the T-DNA copy number. The data suggested that each of three transformants had one T-DNA insert hybridizing to both right- and left-border probes. However, one transformant probably contained four inserts. One DNA fragment of this transformant hybridized to both right- and left-border probes, suggesting that two of the four inserts might be arranged in a directly repeated tandem structure. In addition, both NeoPTase II and PAcTase activities were found in all transformants tested. The neo gene segregation in the F1 progeny of transformants confirmed the low-copy-number T-DNA insertion. Seeds from 10 randomly chosen transformants were germinated on GM containing 50 mg of Km per liter. Two weeks after germination, KmS seedlings had completely etiolated cotyledons, and development of roots and leaves was completely inhibited. By contrast, KmR plants