Fig. 2.

Intermolecular splicing of ribonucleoside-3′-PO₄ terminated DNA. Reaction mixtures (10 µL) containing 50 mM Tris⋅HCl (pH 8.0), 2 mM MnCl₂, 100 µM GTP, 0.5 pmol (0.05 µM) of 5′ ³²P-labeled pD11Rp substrate, 0, 5, or 20 pmol (0, 0.5, or 2 µM) of RtcB as indicated, and either an unlabeled 5′-OH 19-mer RNA acceptor strand (A) or DNA acceptor strand (B) as specified were incubated at 37 °C for 30 min. The products were analyzed by urea-PAGE and visualized by autoradiography. The positions of the substrate, guanylylated intermediate, and ligated product are indicated (Right). The substrate, acceptors, and reaction products are shown (Lower) with the 5′ ³²P-label denoted by •.