Multiple initiation sites of DNA replication flanking the origin region of lambda dv genome.

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Abstract
Early replicative intermediates of .lambda.dv plasmid were prepared by an in vitro replication system in the presence of 2'',3''-dideoxycytidine 5''-triphosphate, an inhibitor of DNA chain elongation. Short-chain DNA produced from regions near the replication origin were purified from the intermediates. A fraction of the DNA was covalently linked to primer RNA. The transition sites from primer RNA to DNA synthesis were mapped along the nucleotide sequence of the genome, by eliminating the RNA by alkaline hydrolysis and labeling the freshly exposed 5'' ends of DNA with 32P. The transition sites were located on both sides of the ori region, which includes four 19-base-pair repeats where one of the .lambda. specific initiator proteins, O, binds. No transition arose within the ori region. The transition sites are multiple on both sides of the ori region and are clustered in one of the 2 strands in such a way that DNA syntheses from the 2 sides converge. The frequency of the leftward DNA synthesis is several times higher than that of rightward synthesis, reflecting the asymmetric bidirectional replication of .lambda.dv DNA.