Cotton is one of the most important fibre crops of India. India is having highest cotton growing area however; its production per unit area is low and stagnant owing to several biotic and abiotic stresses. In order to increase productivity, high yielding, biotic and abiotic stress resistant hybrid/variety need to be developed. Hence, an attempt was made to assess the relationship between heterosis and genetic diversity. Identifying the best genotype combinations for the development of commercial hybrids of cotton remains the main challenge to cotton breeders. Genetic diversity of thirteen parents and heterosis in their forty hybrids which were developed by crossing 8 elite Lines with 5 carefully selected Testers in L × T fashion were used to assess the relationship between heterosis and genetic diversity for eleven important yield and yield contributing traits and three fibre related traits in intra hirsutum cotton (Gossypium hirsutum L.). Diversity studies were conducted by using both Mahalanobis D2 statistic and by random 60 SSR markers, which inferred medium genetic diversity between parents. Correlation coefficient of genetic distance with F1 mean performance, mid, better parent heterosis and specific combining ability for all studied traits were statistically non-significant, indicating that prediction of heterosis for complex traits based on these two genetic diversity estimates is difficult.