<b>Objectives</b><br/>Anopheles sinensis is a predominant malaria vector found throughout the Republic of Korea (ROK). Population genetic analysis can provide insights into the origins and migration patterns of malaria vectors by assessing genetic variation and distribution among populations. <br/><b>Methods</b><br/>In this study, we examined the population genetic structure of An. sinensis in the ROK using mitochondrial cytochrome c oxidase subunit I (COI) marker. <br/><b>Results</b><br/>A total of 903 specimens of An. sinensis were collected from 9 sampling regions, including malaria-endemic and non-endemic areas. All populations displayed characteristics typical of migrant populations, with high haplotype diversity (Hd) and low nucleotide diversity (Pi). Network analysis identified 124 haplotypes grouped into 2 clusters. Both clusters included haplotypes from malaria-endemic and non-endemic areas. Cluster I shared its most recent common ancestry with Chinese reference sequences, while cluster II had related with Japanese reference sequences. Pairwise genetic distance (FST) analysis indicated generally low genetic differentiation among populations. Furthermore, FST values tended to increase proportionally with geographical distance between regions. Analysis of molecular variance confirmed that individual mosquitoes within the population had a large effect on the overall variation. Neutrality tests using 4 methods (Tajima’s D, Fu’s Fs, Fu and Li’s D, and Fu and Li’s F) yielded negative values, suggesting that An. sinensis populations are expanding in all studied regions. <br/><b>Conclusion</b><br/>This study characterizes the genetic attributes of An. sinensis in the ROK, providing valuable insights into the biology of this important malaria vector and contributing useful data for malaria control strategies.