Microalgae play an important function in aquatic environments by serving as the energy foundation of the food chain for all aquatic species. Microalgae produce a wide range of bioproducts, including polysaccharides, lipids, pigments, proteins, vitamins, bioactive chemicals, and antioxidants. In this study, a dataset of identification of three microalgae isolated from freshwater riverine areas at Glami Lemi River, Malaysia is presented. The identification data was acquired using molecular identification using the PCR method and morphological observation. The morphological observation of the microalgae isolates, GL01, GL02, and GL03, showed the closest features and characteristics with Chlorella sp., Ankinstrodesmus sp., and Tetradesmus (Scenedesmus) sp., respectively. The sequence similarity analysis of partial 18s rDNA gene using BLASTn identified the three microalgae species as Chlorella sorokiniana for isolate GL01, Ankistrodesmus fusiformis for isolate GL02, and Tetradesmus obliquus for isolate GL03. Both morphological observation and molecular identification were in agreement in the determination of the species. This is the first report on the three microalgae species isolated from this area.