Magur Clarias batrachus is an indigenous catfish, commonly found in India, Nepal, Bhutan and Bangladesh. Presently, the populations of magur have declined rapidly in their natural habitat mainly due to overexploitation and habitat degradation. Understanding the population genetic structure of the species has significance in improvement of stocks and in conservation of the species. In the present study, simple sequence repeat (SSR) markers were used to differentiate the populations of magur, collected from three geographic locations. For this, a total of 31,814 SSRs were mined from the de novo assembled pooled of whole genome sequence data of C. batrachus. A bioinformatics pipeline with stringent criteria was applied to analyze the data which resulted in selection of 30,142 microsatellite loci falling in the intergenic region. Out of the 25 loci selected for primer development, 16 loci were successfully amplified and 9 loci were found to be polymorphic in this species. The average observed as well as expected heterozygosity in the loci across different stocks varied from 0.652 to 0.688 and 0.864 to 0.873, respectively. These three populations were further segregated into two clusters based on the NJ genetic distance. The Lucknow population formed one cluster, while the Bhubaneswar and Kolkata populations constituted another cluster. A comparable finding was also deduced from the STRUCTURE analyses. The results revealed significant variation among the populations of C. batrachus under study.