The colonization of algal periphyton is a complex process in which algae colonize a new habitat and gradually develop into a mature community. Adhering substrates serve as new habitats and may play an important role in colonization of the periphyton community, but there is no consensus about the colonization process and succession patterns of algal periphyton communities on different substrates. Previous studies have mainly focused on diatoms, which tend to be the first colonizers; little is known about other types of periphytic algae. In addition, most field investigations have placed substrates at the bottom of the water column, and no attention has been paid to the colonization of algae on substrates located at different depths. This study aimed to investigate the process and ecological characteristics of algal periphyton colonization throughout the water column. Two artificial adhering substrates, granite and tile, were placed at the bottom and at mid-depth in a filamentous algae-dominated ecological pond. Algal periphyton on the artificial substrates was sampled every 10 days for analysis of species composition, cell density, and biomass, and the Shannon-Weaver index, Jaccard index, and time to peak biomass of periphytic algae were calculated. Water samples were collected from each site and water quality parameters, including total phosphorous, total nitrogen, nitrate, nitrite, ammonia, and chemical oxygen demand, were determined. The algal periphyton was dominated by diatoms, cyanobacteria, and green algae; 117 species belonging to 73 genera and 8 phyla were identified in the pond. The colonization of algal periphyton on granite and tile showed a similar pattern at both depths. At the early stage of colonization, unicellular diatoms such as Navicula, Fragilaria, and Achnanthes sp. were dominant; the dominant species changed to filamentous algae including Lyngbya, Oscillatoria, and Pseudanabaena sp. at later stage. The type of adhering substrate had no significant effect on species composition, cell density, chlorophyll a, or diversity of algal periphyton. The Jaccard index of the algal communities was similar for granite and tile. On the other hand, the effects of water depth on species composition were significant. There were more periphytic diatom species at the bottom of the water column, while more green algae were present on both substrates placed at mid-depth. The proportion of cyanobacteria gradually increased during colonization. The biomass (chlorophyll a content) of periphytic algae was higher on substrates at the bottom than at mid-depth, but the cell density did not differ significantly. Furthermore, more time was required to reach peak biomass at the bottom. Correlation analyses between cell density and water quality parameters showed that the growth of periphytic algae was mainly dependent on the total phosphorus concentration in the ecological pond.