Figure 3: Schematic overview of the NASBA reaction. Initiation Phase: 1. DNA Primer anneals to RNA template; 2. Primer extension by RT; 3. RNAse H eliminates RNA strand; 4. Primer 2 annealing to DNA template; 5. Polymerase activity of the RT elongates Primer 2, produces double stranded template DNA; 6. T7 RNA polymerase produces RNA transcripts. Amplification Phase steps (1.)-(6.) equal initial phase.