Abstract: A method of detecting rpoB sequences of Mycobacterium tuberculosis present in a biological sample that includes steps of amplifying the M. tuberculosis rpoB sequence in vitro in a nucleic acid amplification mixture that includes specific disclosed primer sequences, and detecting the amplified sequences by using probes that provide information by their specific hybridization to portions of the amplified nucleic acid is disclosed. Compositions for amplifying and detecting in vitro the rpoB sequences of M. tuberculosis in a sample are disclosed.