Figure 2.

Predominant action of Nab2p versus Pab1p in poly(A) tail length control. Polyadenylation kinetics on the pre-cleaved CYC1 precursor (CYC1pre). Two 12-fold reactions were set up and kinetics were initiated by addition of radiolabeled CYC1pre. A 20 µl reaction contains: 30 µg of pab1Δ (YAS394) 3′-end-processing competent yeast cell extract and either 500 ng of recombinant Pab1p (lanes 1–9) or a combination of 500 ng of recombinant Pab1p with 440 ng of recombinant Nab2p (lanes 10–18). Twenty microliters from each reaction mixture were withdrawn at the indicated time points, and the RNA content was extracted, separated on a 6% polyacrylamide-8.3 M urea denaturing gel and visualized by autoradiography. M, molecular weight markers were as described in Figure 1.