Tor tor, commonly known as the tor mahseer or 'deep bodied' mahseer, is a cyprinid fish species found in fast-flowing rivers and streams with rocky bottoms in Bangladesh, Bhutan, India, Nepal, and Pakistan. It is a well known commercially important food and game fish. Its population is rapidly declining in its native range due to overfishing. Recently, the species is assessed as data deficient under IUCN (2018) due to ambiguity in taxonomic identification and distribution. Moreover, there is meagre knowledge about genetic structure of the species which is necessary for planning management,conservation and stock assessment of this species. Therefore, microsatellite markers were used to assess the genetic variation in this species. A total of twenty polymorphic single-locus microsatellite loci were developed through next generation sequencing technology, to genotype n=140 samples from four distant riverine populations of India, namely, Penganga, Narmada, Madar and Godavari, for determining intra-specific genetic variation in golden mahseer, T. tor. The mean number of observed alleles per locus ranged from 1.65 to 2.30. The observed and expected heterozygosities ranged from 0.2256 to 0.3834 and from 0.1690 to 0.3537, respectively. The pair wise FST value ranges from 0.09679(P=0.0000) to 0.29122(P 0.001). Significant genotype heterogeneity (P<0.001) and FST =0.182 (P=0.0000) over all loci indicated that the samples are not drawn from the same gene pool. The microsatellite loci exhibited high level of polymorphism with PIC values ranging from 0.067 to 0.553. The identified microsatellite loci are promising for use in fine scale population structure analysis of T. tor . No significant linkage disequilibrium was detected among the loci. Five loci showed significant deviation from Hardy-Weinberg equilibrium, but none of them had evidence for null alleles. These microsatellite loci will be useful for genetic diversity studies and conservation of golden mahseer and its closely related species.