The complete cDNA sequences of Aquaporin 3 (AQP3) from gills of Sarotherodon melanothern was cloned using rapid amplification of cDNA ends (RACE). The full length of AQP3 cDNA was 1894 bp, containing 912 bp open reading frame and encoding 303 amino acids, with a 98 bp 5’ untranslated regions (UTR) and 884 bp 3’-UTR. The amino acid sequence comparison showed that AQP3 of S. melanothern shared the highest identity with Oreochromis mossambicus (94%). Relative tissue expression of AQP3 mRNA under different salinity stresses were estimated by real-time quantitative PCR. The higher expression were detected in skin, gill and muscle than in other tissues at 0 and 15 salinity, moreover, the expression level in 15 salinity was lower than that in freshwater in all tissues. The expression level was higher in intestine than in other tissues at 30 salinity. The results suggested S. melanothern could utilize AQP3 in different tissue to respond to water transfer under different osmoregulation process.