Genetic similarity and diversity of catfish Clarias batrachus (Linn.1758) populations collected from three regions of Indian riverine system were examined using randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR). Out of 22 random primers tested, six primers produced 462 RAPD bands ranging from 105 to 128 polymorphic bands per primer in size between 100 and 1,200 bp. The polymorphic bands in these populations ranged from 26.5 to 30.5. Polymorphic bands within populations ranged from 25 to 35.7%. The degree of similarity within Hussainabad population varied from 0.125 to 0.842 based on average level of band sharing (BS) values. The level of band sharing values within the catfish populations were 0.26 +/- 0.021 for Banaras, 0.60 +/- 0.033 for Bhubaneshwar and 0.377 +/- 0.058 for Hussainabad respectively. The results from the present study indicate that there is great degree of genetic similarity between Bhubaneshwar and Hussainabad populations where as Banaras catfish population is distinct. It may appear that Bhubaneshwar and Hussainabad are geographically connected by rivers and most of the major catfish hatcheries are located in this region, therefore the individuals from these populations are get reared in the same environmental conditions, migration or by inbreeding during several generations may be possible. This may be the reason that catfish population is lacking genetic diversity in major riverine system of India. In nearer future, the lack in genetic diversity can lead to inbreeding which can be resulted in poor growth and disease susceptibility, Bhubaneshwar and Hussainabad catfish population may have this problem.