The barbels of the subfamilies ´Poropuntinae´ and Smiliogastrinae within the family Cyprinidae play a significant role as a food source for fish in artisanal fisheries and are highly valued as ornamental fish in Thailand. In this study, we employed both conventional and molecular cytogenetics to analyze the karyotype of 15 fish species from two cyprinid lineages. All analyzed species had a diploid chromosome number of 2n = 50. Despite sharing the same 2n, our analyses revealed species-specific distribution patterns of the mapped microsatellite motifs [(CA)₁₅, (TA)₁₅, (CAC)₁₀, and (CGG)₁₀]. They were predominantly found at telomeric sites of all-to-few chromosomes. Additionally, some species exhibited a widespread distribution of the mapped microsatellites across the chromosomes while others showed no signal. These variations reflect the evolutionary divergence and chromosomal diversity within the cyprinids. Thus, our findings support the 2n stability in cyprinoid lineages while emphasizing the intrachromosomal evolutionary diversity accompanied by species-specific microsatellite distribution.