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1Division of Avian Genetics and Breeding, Molecular Genetics Laboratory, ICAR-Central Avian Research Institute, Izatnagar, India 2Division of Avian Genetics and Breeding, Molecular Genetics Laboratory, ICAR-Central Avian Research Institute, Izatnagar, India 3Division of Animal Genetic and Breeding, Veterinary College and Research Institute, Orathanadu (Tamil Nadu), India  RAHIM, A., S. KUMAR, R. YADAV, J. DEBNATH, J. KRISHNAN: Genetic variability determination in a long-term selected Rhode Island Red chicken strain using microsatellite markers. Vet. arhiv 87, 511-522, 2017. ABSTRACT Genetic improvement needs the existence and precise estimation of genetic variability, and microsatellites are a molecular marker of choice for its assessment. Rhode Island Red (RIR) is a brown-egger chicken breed. A RIR population, selected on the basis of 40-week part-period egg production for 30 generations, maintained as a closed flock to develop multi-colored strains for backyard farming, were studied to determine genetic variability and heterozygosity using microsatellite markers. Genomic DNA from 76 randomly selected birds was analyzed at 10 microsatellite loci, alleles were separated on 3.4% MetaPhor Agarose, sized using Quantity-One software and analyzed by POPGENE v1.32. Nine loci demonstrated polymorphism resolving 30 alleles, the average number of alleles/ locus being 3.00 ± 1.41. Allele numbers varied from two to six and size ranged from 102 to 320bp. The average polymorphic information content and Nei’s heterozygosity were 0.3134 ± 0.064 and 0.4119 ± 0.2575, respectively, suggesting these loci to be moderately polymorphic and informative. The observed (Na) and effective number (Ne) of alleles and Shannon’s index averaged 3.0000 ± 1.4142, 2.0324 ± 0.9416 and 0.7342 ± 0.4649, respectively. Ne was less than Na at most loci, indicating the prevalence of heterozygosity at these loci. Mean expected heterozygosity was higher than mean observed heterozygosity at most loci, indicating that the population was not in Hardy-Weinberg equilibrium but was under the influence of some forces such as selection, etc. The same was also demonstrated by Chi-square and G-square tests. It may be inferred that long-term selection for 40-week part-period egg production has resulted in Hardy-Weinberg disequilibrium at the studied microsatellite loci, thereby suggesting a probable association between these microsatellite loci and layer economic traits in RIR chicken, and this might be useful in marker assisted selection for egg production in future.