Identification of Six Single-Strand Initiation (ssi) Signals for Priming of DNA Replication in Various Plasmids

Abstract

Using a mutant M13 phage derivative lacking a great part of the complementary strand synthesis origin, we identified six single-strand initiation (ssi) signals for DNA replication in pACYC184, pLG214, pGKV21, and pDPT270 plasmids, and named them tex:\small\textstyle$ssiA_{YC}$, tex:\small\textstyle$ssiA_{LG}$, tex:\small\textstyle$ssiB_{LG}$, tex:\small\textstyle$ssiA_{KV}$, tex:\small\textstyle$ssiA_{PT}$, and tex:\small\textstyle$ssiB_{PT}$, respectively. Two of them were from pDPT270, one from downstream the on of pACYC184, two from pLG214, one from upstream the plus origin of pGKV21. Introduction of these ssi signals into the deleted tex:\small\textstyle$ori_c$ site of a mutant filamentous M13 phage (tex:\small\textstyle$M13{\Delta}lac182$) resulted in the restoration of growth activity of this phage. These ssi signals were classified into a number of groups on the basis of sequence similarity. tex:\small\textstyle$ssiA_{YC}$ and tex:\small\textstyle$ssiA_{LG}$ show extensive sequence homology to the n'-site (primosome assembly sites) of ColE1, whereas tex:\small\textstyle$ssiB_{PT}$ is homologous to the n'-site of tex:\small\textstyle${\Phi}X174$. tex:\small\textstyle$ssiA_{PT}$ belongs to G4-type ssi signals which require only dnaG primase and SSB protein for the priming of replication. In addition, possible biological roles of these ssi signals are discussed.