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Separation and Partial Characterization of Two Ribonucleic Acid Polymerases from Pea Seedlings ¹

^a Department of Botany and the Institute of Plant Development, University of Wisconsin, Madison, Wisconsin 53706

Two DNA-dependent RNA polymerases (ribonucleoside triphosphate:RNA nucleotidyl transferase, EC 2.7.7.6) have been isolated from pea (Pisum sativum) seedlings. The enzymes were solubilized by sonication in high salt buffer and were separated by chromatography on diethylaminoethyl cellulose using a linear salt gradient. Polymerase I eluted at 0.10 M (NH₄)₂SO₄, accounted for about 10% of the recovered activity and was completely insensitive to -amanitin. Polymerase II eluted at 0.14 M (NH₄)₂SO₄, accounted for the remaining 90% of recovered activity and was strongly inhibited by -amanitin. Both enzymes preferred denatured to native DNA as template, both showed an absolute requirement of divalent cation, and both were sensitive to the ionic strength of the assay medium. The developing pea seedling seems a promising system for studies of possible changes in relative activities and roles of multiple RNA polymerases during eukaryotic development.