The Nilakottai ecotype, a renowned indigenous variety of tuberose cultivated by farmers in the Dindigul district, is distinguished by its unique fragrance. To further enhance the floral characteristics of this tuberose ecotype, mutation breeding was undertaken at Tamil Nadu Agricultural University, Coimbatore. The present study involved 11 treatments, each replicated 3 times, using a CRD (completely randomized design). The results revealed that the LD50 for the Nilakottai ecotype with ethyl methane sulphonate (EMS) was 0.42 %. As the EMS concentration increased, there was a reduction in both sprouting and survival, with a 38.2 % and 56.24 % decrease respectively, at the highest concentration. Morphological parameters such as root length, shoot length, leaf length, leaf width and the number of leaves also showed a decrease of 78.80 %, 71.16 %, 87.29 %, 61.90 % and 62.99 % respectively, compared to the control at 1.0 % EMS. Pollen viability studies indicated a reduction in viability with increasing EMS concentration, showing 85.74 % viability in the control group and only 31.54 % at 1.0 % EMS. Analysis of antioxidant enzymes revealed a significant increase in catalase (CAT), peroxidase (POD) and superoxide dismutase (SOD) levels with higher EMS concentrations. Overall, EMS had a pronounced effect on various traits, including germination, growth and antioxidant activity.