Introduction: Tuberculosis is one of the oldest diseases with high morbidity and mortality. India accounts for one– fourth of the global TB burden (2.7million cases). Material and methods: A total 1056 patients were included in this study. All patients were requested to give two sputum samples, spot sputum sample (at the time of visit) and early morning sputum for mycobacterial testing. All spot samples tested with smear microscopy and Xpert MTB/Rif assay. Rifampicin resistant samples compared with conventional method. Results: Out of 1056 sputum samples GeneXpert MTB Rif assay detected positive 204 (19.3%), not detected 827 (78.31%). There is significant difference founded for positivity in spot samples and early morning sputum samples (P =0.0026). Rifampicins resistant were 8 (3.9%) andshown 100% sensitivity, specificity with Conventional method. Conclusion: For diagnosing tuberculosis and detecting Rifampicin resistance GeneXpert MTB/RIF has been especially recommended.