The DUF1644 family is a highly conserved and functionally unknown protein family and has been demonstrated significantly improved the salt tolerance of rice (nonhalophytes). However, little is understood about the function of the DUF1644 gene in halophytes. Further studies are required to determine whether it has a similar function in halophyte species. In our study, the functions of ZmDUF1644 from halophyte Zoysia matrella Z123 species were investigated under salt stress. Phylogenetic analysis clustered ZmDUF1644 and OsSDIP361 in the same group, which function as a salttolerant genes in rice. Our experimental data suggested that the ZmDUF1644 gene significantly alleviated the growth inhibition of transgenic Arabidopsis thaliana seedlings caused by salt stress. The maximum root length, fresh weight, and photosynthetic pigment content of transgenic seedlings were obviously better than that of the wild type. Moreover, a relatively lower relative electrolyte leakage rate, toxic ion (Na+) content, and Na+/K+ ratios were also found in the transgenic line when compared with the wild type under salt stress. In conclusion, this study revealed that the ZmDUF1644 gene was overexpressed in Arabidopsis thaliana significantly enhancing the salt tolerance of seedlings. It is the first report on the functions of ZmDUF1644 from halophyte Zoysia matrella which has a similar function in halophyte species.