Figure 5.

Differential polyadenylation of pppRNA I and pRNA I. Universally ³²P-labeled pppRNA I (1.2 pmol) and 5′ end kinase-labeled pRNA (1.2 pmol) were used as substrates. M, 1-kb DNA marker (Life Technologies), which was labeled with [α-³²P]dATP by using Klenow fragment of E. coli DNA polymerase I. The rate of initiation of poly(A) tails (solid lines) and the average length of poly(A) tails (dashed lines) are shown for pppRNA I (●) and pRNA I (■). Reaction conditions were as described in Fig. 1, except that 70 fmol of PAP I was used and an extra time point (32 min) was taken. The times required for adenylation of 50% of the substrate were 2.2 ± 0.1 min for pppRNA I and 0.4 ± 0.1 min for pRNA I. The average rate of poly(A) tail growth, which was defined as the average length of the poly(A) tail at each time point divided by the number of minutes, was 3.75 nt/min for pppRNA I and 11.68 nt/min for pRNA I.