Eighty-two varieties of rice from different regions in Thailand were selected to explore the Waxy (Wx)gene diversity and indica-japonica differentiation of chloroplast DNA. A comparison of the 5' splice site in the first intron was made between glutinous and nonglutinous rice. It revealed that non-glutinous with low-amylose content and glutinous rice were characterized as the W x ^b allele based on the G-to-T base substitution, whereas non-glutinous rice with intermediate and high amylose carried the Wx ^a allele. Four Wx microsatellite alleles, (CT) _n repeat, (n = 16,17,18 and 19) were found in glutinous rice. In contrast, non-glutinous rice showed five Wx microsatellite alleles (n = 11, 16, 17, 18 and 19). The (CT)₁₇ allele was prominent allele in Thai population, while the (CT)₁₁ allele was found only in intermediate and high amylose rice varieties from southern Thailand. Almost all of upland rice grown by various ethnic groups in northern Thailand were characterized as japonica type based on their having the PstI-12 fragment in their cpDNA, whereas most of rainfed lowland varieties from other regions of Thailand were indica. This exploration of DNA-based genetic markers is important, as it enhances our ability to describe and manipulate sources of genetic variation for rice breeding programs.