Two deep-coverage bacterial artificial chromosome (BAC) libraries of Oryza sativa japonica cv. Tainung 67 (TNG 67), a popular genetic stock in breeding programs and scientific research in Taiwan, have been constructed to facilitate positional cloning of rice genes and to analyze variety-specific genome composition, toward rice structural and functional genomic studies. Good high-molecular-weight DNA was produced well by nuclei preparation from two- to three-week-old seedlings, partially digested by either HindIII or EcoRI, and two cycles of size selection by pulsed-field gel electrophoresis (PFGE). The HindIII library consists of 45,312 clones, near lack of false positive clones, with an estimated average insert size of 138.4 kb and coverage of 15.1× haploid genome equivalents. The EcoRI library consists of 9984 clones, 2% of false positive clones, with an estimated average insert size of 137.8 kb and coverage of 3.2× haploid genome equivalents. Nine single-copy sequence tagged site (STS) markers, located on different chromosomes, were used to screen the two libraries, which a single BAC clone double-spotted on three filters, and hybridized 16-29, with an average of 21.2 BACs. As a result, these two libraries can complement each other and cover nearly 100% of the rice genome, consequently providing efficient tools to isolate any genes of interest. Resources, high-density filters, individual clones, and whole libraries, are available for public distribution and may be accessed at the Institute of Plant and Microbial Biology, Academia Sinica, or Arizona Genomic Institute (AGI).