Genetic diversity of six populations of bartail flathead (Platycephalus indicus Linnaeus, 1758) was investigated using amplified fragment length polymorphism (AFLP). A total of 118 reproducible bands amplified with ten AFLP primer combinations were obtained from 42 fishes that were collected from six different locations in the northern part of the Persian Gulf. The percentage of polymorphic bands was 57.06%. Nei’s genetic diversity was 0.1808±0.2074 - 0.2164±0.1903, and Shannon’s index was 0.2675±0.2949 - 0.3280±0.2727. The results of AMOVA analysis indicated that 66% and 34% of the genetic variation occurred within and among the populations and gene flow was 0.6454. The estimated level of population differentiation as measured by average Fst value across all loci was 0.327. Analyses revealed significant genetic differentiation with low gene flow among the six locations, indicating at least six separate populations of bartail flathead in the northern part of the Persian Gulf.