Initiation sites for discontinuous DNA synthesis of bacteriophage T7.

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Abstract
The discontinuous replication of bacteriophage T7 DNA is primed by tetraribonucleotides (major component) or pentaribonucleotides. Both tetramers and pentamers start with pppA-C and are rich in A and C at the 3rd and 4th nucleotides. The sites of transition from primer RNA to DNA in vivo were located on a 340 nucleotide segment of the H strand of the T7 genome by 32P-labeling in vitro of the 5''-hydroxyl ends of DNA resulting from alkaline hydrolysis of RNA-linked T7 DNA fragments. Five strong transition sites were detected with a common sequence 5''-G-A-C-N1-N2-N3-N4-3'', in which N1 was either C or A, N2 was A, C or G and either N3 or N4 was the nucleotide for the switchover to DNA synthesis. The complementary sequence 3''-C-T-G-G/T-N''2-(N''3)-5'' in the template strand is probably the most frequently used signal for synthesis of primer RNA. Primer-RNA synthesis starts at a precisely defined nucleotide, but the transition to DNA synthesis varies within 2 nucleotides. Because the observed signal sequence would be present on a statistical basis once per 128 nucleotides, only about 10% of the existing signals are used for primer synthesis in each round of replication so that nascent fragments 1000-2000 long result. This provides an unexpected flexibility for RNA priming of DNA synthesis.