Brown planthopper (BPH) is a destructive insect pest of rice in Asia. Identification and the incorporation of new BPH resistance genes into modern rice cultivars are important breeding strategies to control the damage caused by new biotypes of BPH. In this study, a major resistance gene, Bph18(t), has been identified in an introgression line (IR65482-7-216-1-2) that has inherited the gene from the wild species Oryza australiensis. Genetic analysis revealed the dominant nature of the Bph18(t) gene and identified it as non-allelic to another gene, Bph10 that was earlier introgressed from O. australiensis. After linkage analysis using MapMaker followed by single-locus ANOVA on quantitatively expressed resistance levels of the progenies from an F2 mapping population identified with marker allele types, the Bph18(t) gene was initially located on the subterminal region of the long arm of chromosome 12 flanked by the SSR marker RM463 and the STS marker S15552. The corresponding physical region was identified in the Nipponbare genome pseudomolecule 3 through electronic chromosome landing (e-landing), in which 15 BAC clones covered 1.612 Mb. Eleven DNA markers tagging the BAC clones were used to construct a high-resolution genetic map of the target region. The Bph18(t) locus was further localized within a 0.843-Mb physical interval that includes three BAC clones between the markers R10289S and RM6869 by means of single-locus ANOVA of resistance levels of mapping population and marker-gene association analysis on 86 susceptible F2 progenies based on six time-point phenotyping. Using gene annotation information of TIGR, a putative resistance gene was identified in the BAC clone OSJNBa0028L05 and the sequence information was used to generate STS marker 7312.T4A. The marker allele of 1,078 bp completely co-segregated with the BPH resistance phenotype. STS marker 7312.T4A was validated using BC2F2 progenies derived from two temperate japonica backgrounds. Some 97 resistant BC2F2 individuals out of 433 screened completely co-segregated with the resistance-specific marker allele (1,078 bp) in either homozygous or heterozygous state. This further confirmed a major gene-controlled resistance to the BPH biotype of Korea. Identification of Bph18(t) enlarges the BPH resistance gene pool to help develop improved rice cultivars, and the PCR marker (7312.T4A) for the Bph18(t) gene should be readily applicable for marker-assisted selection (MAS).