Tonoplast intrinsic proteins (TIPs), which comprise a subfamily of aquaporin (AQP) mediating the water balance within plant cells, could be divided into five phylogenetic groups on the basis of sequence similarity, i.e., TIP1–TIP5. Based on the full-length transcriptome previously described in tigernut (Cyperus esculentus), a tuber-abundant TIP gene named CeTIP4;1 was cloned and characterized in this study. Sequence analyses revealed that CeTIP4;1 contained two conservative introns with a complete coding sequence of 753 base pairs (bp), putatively encoding 250 amino acids with the theoretical molecular weight of 25.57 kDa, the isoelectric point (pI) value of 6.02, and the grand average of hydropathicity (GRAVY) value of 0.884, implying its acidic and hydrophobic features. The deduced peptide possessed one conservative major intrinsic protein (MIP) domain and exhibited similar structural features with the structure resolved AtTIP2;1. Classification of CeTIP4;1 into the TIP4 group was supported by the phylogenetic analysis, which showed that CeTIP4;1 clusters with AtTIP4;1, OsTIP4;1, OsTIP4;2 and OsTIP4;3 with sequence similarities of 69.62%–82.94%. After gene cloning, a plant expression vector for subcellular localization, i.e., pNC-Cam1304-CeTIP4;1, was successfully constructed. When transiently overexpressed in tobacco (Nicotiana benthamiana) leaves, confocal laser scanning microscopy imaging showed that CeTIP4;1 was unexpectedly localized to the cell membrane. Further expression analysis indicated that CeTIP4;1 exhibited an apparent tissue-specific expression patter, predominately expressed in tuber, root, rhizome, and shoot apex. Moreover, among the four typical stages of developmental tuber examined in this study, i.e., initiation, medium swelling, late swelling, and maturation, CeTIP4;1 exhibited a bell-like expression pattern, peaking in medium swelling tubers and lowest in maturation tubers. These results lay a solid foundation for further uncovering the mechanism of water balance in tigernut tubers.