Figure 3.

Assaying for intrinsic transcript termination at T₇Te on ssDNA. C₄₆ complexes bound to Ni²⁺-NTA agarose in TGK-B₄₀M₄ were chased with 500 μM ATP, 500 μM GTP, 500 μM CTP, and 500 μM UTP for 10 min at 37°C in the presence of rifampicin at 20 μg/ml and yeast Torula RNA at 0.8 mg/ml. The lanes are assigned as for Fig. 2A. The T₇Te terminator is at positions +95 and +96, whereas transcription to the end of the DNA template generates a run-off RNA of +146 nucleotides. C₄₆ complexes in lanes 5–8 were digested with Exo III at 5,000 units/ml for 5 min at 37°C. Unlike C₄₇, some of the C₄₆ complexes failed to resume elongation after treatment with Exo III (see lanes 6 and 7). The mechanism of this inactivation is unknown but similar observations have been made by others (29, 30).