qCTS-9 is reported to be a rice cold-tolerant gene, which functions stably under a wide range of low temperature conditions at seedling stage. Detailed knowledge of the haplotype and adaptive evolution pattern of qCTS-9 will provide support for breeding cold-tolerant rice varieties. In this study, 116 accessions of cultivated rice (Oryza sativa L.) and 37 accessions of wild rice (Oryza rufipogon Griff.) were used to analyze the nucleotide diversity and haplotypes of qCTS-9. There are a total of 14 SNPs in the coding region of qCTS-9, and these SNPs comprised eight haplotypes. Four Non-synonymous SNPs constituted three functional haplotypes, which showed significant differences in cold tolerance. The SNP at the 1535 bp in the coding region was hypothesized to be the key mutation site of qCTS-9. The analysis of cis-acting elements in the promoter region of qCTS-9 showed that a SNP (G→A) at -1107 bp resulted in the deletion of a MYB recognition site in the cold-sensitive indica haplotype Hap_4, which may lead to a decrease in cold tolerance in indica rice.Wild rice contains most of the variations of qCTS-9 gene in cultivated rice. During the differentiation from wild rice to indica and japonica rice, cold-tolerant related alleles or mutations were fixed and expanded, which in turn enhanced its regional adaptability.