Upland rice is one of the most important ecotypes growing on dry or rain-fed land and exhibiting better nitrate utilization. However, the differences in nitrogen uptake and the underlying molecular mechanisms between upland and lowland rice remain unknown. In this study, a nitrate transporter gene, OsNPF7.1, was identified by analyzing nitrogen-related gene differences in the genomic and transcriptomic profiles of the two ecotypes within the japonica subspecies. OsNPF7.1 is mainly expressed in the endodermis, xylem parenchyma cells and phloem parenchyma cells and is directly activated by OsNAC42. Ectopic expression of OsNPF7.1 in Xenopus oocytes showed that the gene functions as both a low- and high-affinity NO 3 - transporter. Nitrate uptake and redistribution were impaired in npf7.1 mutants. Grain yields and nitrogen use efficiency (NUE) were elevated in OsNPF7.1 overexpression transgenic plants grown under both low and normal nitrogen supply conditions. Moreover, variation in the putative promoter increases OsNPF7.1 expression and affects nitrate acquisition and redistribution in rice. The potential elite haplotype of OsNPF7.1 in upland rice could be beneficial for breeding rice varieties with high NUE.