PRIMER-DNA FORMATION DURING SIMIAN-VIRUS 40 DNA-REPLICATION INVITRO

Studies of simian virus 40 (SV40) DNA replication in vitro have identi fied a small (approximately 30-nucleotide) RNA-DNA hybrid species term ed primer-DNA. Initial experiments indicated that T antigen and the po lymerase alpha-primase complex are required to form primer-DNA. Prolif erating cell nuclear antigen, and presumably proliferating cell nuclea r antigen-dependent polymerases, is not needed to form this species. H erein, we present an investigation of the stages at which primer-DNA f unctions during SV40 DNA replication in vitro. Hybridization studies i ndicate that primer-DNA is initially formed in the origin region and i s subsequently synthesized in regions distal to the origin. At all tim e points, primer-DNA is synthesized from templates for lagging-strand DNA replication. These studies indicate that primer-DNA functions duri ng both initiation and elongation stages of SV40 DNA synthesis. Result s of additional experiments suggesting a precursor-product relationshi p between formation of primer-DNA and Okazaki fragments are presented.