In this study, mitochondrial DNA (mtDNA) analysis was carried out on 9 Bronze Age horses recovered from Dashanqian and Jinggouzi archaeological sites in Chifeng region, Inner Mongolia. China to explore the origin of Chinese domestic horses. Both mtDNA 16S rRNA gene and control region (D-loop) fragments of ancient horses were amplified and sequenced. The analysis of the highly conservative 16S rRNA gene sequences indicated that the burial environment of Chifeng region is suitable for the preservation of ancient DNA (aDNA). Combing 465 mtDNA D-loop sequences representing different breeds from East Asia, Central Asia, Near East and Europe, we constructed a phylogenetic network to investigate the relationship between ancient and modern horses. The phylogenetic network showed that the 9 horses were distributed into different modern horse clusters which were closely related to them representing a certain geographical distribution. Our results showed that the maternal genetic line of the ancient horses in Chifeng region was highly diversified, which contributed to the gene pool of modern domestic horses and suggested a complex origin of domestic horses in China. In this study, mitochondrial DNA (mtDNA) analysis was carried out on 9 Bronze Age horses recovered from Dashanqian and Jinggouzi archaeological sites in Chifeng region, Inner Mongolia. China to explore the origin of Chinese domestic horses. Both mtDNA 16S rRNA gene and control region The analysis of the highly conservative 16S rRNA gene sequences indicated that the burial environment of Chifeng region is suitable for the preservation of ancient DNA (aDNA). Combing 465 mtDNA D-loop (D-loop) fragments of ancient horses were amplified and sequenced. The sequences representing different breeds from East Asia, Central Asia, Near East and Europe, we constructed a phylogenetic network to investigate the relationship between ancient and modern horses. The phylogenetic network showed that the 9 horses were distributed into different modern horse clusters which were closely related to them representing a certain geographical distribution. Our results showed that the maternal genetic line of the anci ent horses in Chifeng region was highly diversified, which contributed to the gene pool of modern domestic horses and suggested a complex origin of domestic horses in China.