Microbial resistance to existing antibiotics has led to an increase in the use of medicinal plants that show beneficial effects for various infectious diseases. To evaluate the susceptibility of mycobacteria to Nigella sativa extract. In the present study Nigella sativa was extracted with methanol, cholorform and water using Soxhlet extractor. The extracts were than tested against Mycobacterium sp. using mycobacteriophage propagation. The Minimal Inhibitory Concentration (MIC) of N. sativa seeds were determined against all three mycobacterial strains using Luciferase Reporter Phage (LRP) assay. The TLC profiling of N. sativa seeds was carried out. In this study, the methanolic and water extract of N. sativa seeds showed inhibition against M. tuberculosis H37Rv, All drug Sensitive M. tuberculosis, MDR M. tuberculosis respectively. The methanol extract showed least inhibition at concentration of 50 µg/ml, 250 µg/ml and 100 µg/ml against M. tuberculosis H37Rv, All drug sensitive M. tuberculosis and MDR M. tuberculosis respectively. N. sativa seed is a traditional medicine, which possesses anti-tubercular property. This plant has great potential to develop as a natural drug for the treatment of tuberculosis including drug resistant TB.