Nile tilapia (Oreochromis niloticus L.) is indigenous species to Ethiopia and constitutes major proportion in the country’s fish production. In an attempt to select better performing strains for aquaculture development, tilapia populations from different Ethiopian rift valley lakes showed different growth performances in pond culture. Investigation of desired culture characteristics of target tilapia populations is required to improve their productivity in aquaculture system. The current study was made to investigate phenotypic characters of the tilapia populations in three geographically isolated Ethiopian rift valley lakes (Chamo, Koka and Ziway). A total of 450 adult tilapias of commercial catches were sampled from the three lakes and their phenotypic characters were analyzed during May 2018 to March 2019. Twenty six morphometric character indices, eight meristic counts, total length, standard length, total weight, length-weight relationship and Fulton’s condition factor were considered in the analysis. The results revealed significant differences (p ≤ 0.05) in most of the morphometric character indices, meristic counts, mean length and weight and Fulton’s condition factor among the three tilapia populations. Chamo tilapia population were found to have highest mean values of total weight, total length and standard length while Koka population have highest mean value of Fulton’s condition factor and positive allometric growth as characters desired in aquaculture. Further research is required to investigate whether the fish populations could maintain those characters in pond cultures.