a Self Transmissible, Narrow-Host-Range Endogenous Plasmid of Rhodobacter sphaeroides 2.4.1 : Physical Structure, Incompatibility Determinants, Origin of Replication, and Transfer Functions

Abstract

Rl~todobacler sphaeroides 2.4.1 naturally harbors live cryptic endogenous ltlasrnids (C. S. Fornari, Rl. M'atlkins, and S. Kaplan, Plasr~id1 1:39-47, 1984). The s~nallestp las~iiid( pRS24le), with a ~iioleculars ize of 42 kb, was observed to be a self-transmissible plasmid which can transfer only to certain strains of K. sphaqeroides. Transfer frequencies can be as high as lo-' to lo--' per donor under optinial mating conditions in liquid media in the absence ofoxygen. pKS241e, designated the S factor, was also shown to possess a narrow host range, failing either to re~llicateo r to be maintained in Eschericl~iac oli, AgroDacferircrr~~ trmcfacierrsa, nd Hl~iz~~~bni~nemlil otiI. t was furtller revealed that one of the remaining four endogenou? plasmids, pliSZlld, was z!so transr~iissible at a frequency similar to that of the S factor. As a cointegratc with pSUP203, S was maintained in E. coli, provid~ng sufficient DNA froni wliicl: a physical ni;~p of S could be constructed. I'rogressivc subcloning of S-faclor DNA, in conjunction wit11 assays of plasmid transfer, led to the locali~atiori and identification of oriV (IncA , IncB, and the putative orir locus. 'She DNA sequence of the 427 bp containing oril;, revealed topological similarity to other described oriT sequences, consisting of an A-T-rich DNA region, several direct and inverted repcbats, and putative integration host factor (1HF)-binding sites, and was shown to be fu~nctional in promoting pla imid transfer.