Plus-origin mapping of single-stranded DNA plasmid pE194 and nick site homologies with other plasmids.
Staphylococcus aureus plasmid pE194 manifests a natural thermosensitivity for replication and can be established in several species, both gram positive and gram negative, thus making it attractive for use as a delivery vector. Like most characterized plasmids of gram-positive bacteria, pE194 generates single-stranded DNA. The direction of pE194 replication is clockwise, as determined by the strandedness of free single-stranded DNA. Significant homology exists between a 50-base-pair sequence in the origin of pE194 and sequences present in plasmids pMV158 (Streptococcus agalactiae), pADB201 (Mycoplasma mycoides), and pSH71 (Lactococcus lactis). We used an initiation-termination reaction, in which pE194 initiates replication at its own origin and is induced to terminate at the related pMV158 sequence, to demonstrate that pE194 replicates by a rolling-circle mechanism; the initiation nick site was localized to an 8-base-pair sequence.
Institut de Biotechnologie, Institut National de la Recherche Agronomique--Domaine de Vilvert, Jouy en Josas, France.
S
S
Sozhamannan
P
P
Dabert
V
V
Moretto
S D
SD
Ehrlich
A
A
Gruss
D001483:Base Sequence; D003001:Cloning, Molecular; D004261:DNA Replication; D004277:DNA, Single-Stranded / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D008969:Molecular Sequence Data; D010957:Plasmids; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D013211:Staphylococcus aureus / Q000235:genetics*; D013728:Terminator Regions, Genetic
D004277:DNA, Single-Stranded