Identification of a toxin coding fragment in pBSSB1, a linear plasmid from Salmonella enterica serovar Typhi that can stabilize a multicopy plasmid

Objective: To identify the region conferring stability to pBSSB2 (a linear plasmid, pBSSB1, containing a kanamycin cassette), which is unique to Indonesian isolates of Salmonella enterica serovar Typhi. Methods: The open reading frame (ORF) 009 was identified as a toxin coding gene in the plasmid through introduction of translational termination codons in the ORF. Results: The stability function was located in a fragment that spanned nucleotides 5 766 to 6 828 in the linear plasmid genetic map. Ectopic expression of ORF009 in pBAD18 vector indicated ORF009 codes for a toxin. This fragment could stabilize plasmid pUC18 previously destabilized through mutation of the pcnB (plasmid copy number control) gene that codes for polyA polymerase. Majority of the cells expressing ORF009 were non-viable according to phase contrast microscopy. Conclusions: This study demonstrated that a linear plasmid fragment that carries a gene encoding a toxin possibly conferred stability to the parent plasmid. It was able to stabilize a multicopy plasmid of Escherichia coli.

Identification of a toxin coding fragment in pBSSB1, a linear plasmid from Salmonella enterica serovar Typhi that can stabilize a multicopy plasmid

Objective: To identify the region conferring stability to pBSSB2 (a linear plasmid, pBSSB1, containing a kanamycin cassette), which is unique to Indonesian isolates of Salmonella enterica serovar Typhi. Methods: The open reading frame (ORF) 009 was identified as a toxin coding gene in the plasmid through introduction of translational termination codons in the ORF. Results: The stability function was located in a fragment that spanned nucleotides 5766 to 6828 in the linear plasmid genetic map. Ectopic expression of ORF009 in pBAD18 vector indicated ORF009 codes for a toxin. This fragment could stabilize plasmid pUC18 previously destabilized through mutation of the pcnB (plasmid copy number control) gene that codes for polyA polymerase. Majority of the cells expressing ORF009 were non-viable according to phase contrast microscopy. Conclusions: This study demonstrated that a linear plasmid fragment that carries a gene encoding a toxin possibly conferred stability to the parent plasmid. It was able to stabilize a multicopy plasmid of Escherichia coli.

Characterization of The Essential Gene Components for Conjugal Transfer of Streptomyces lividans Linear Plasmid SLP2 / 生物化学与生物物理进展

Commonly, the interbacterial transfer of circular plasmids is initiated by nicking at an internal sequence, oriT, followed by transferring one strand as single-stranded DNA through a type Ⅳ secretion channel on cell membrane. In contrast, Streptomyces conjugative linear plasmids, containing a free 3'-end but a protein-capped 5'-end, can potentially undergo cell-to-cell transfer by transfer of non-nicked DNA. It was reported that circular derivatives of the Streptomyces lividans linear plasmid SLP2, as well as the parental linear plasmid itself can transfer efficiently. And the genetic requirements for such transfer was described. Efficient transfer of plasmid requires six co-transcribed SLP2 genes, encoding a Tra-like DNA translocase, cell wall hydrolase, two cell membrane proteins that interact with an ATP binding protein, and a protein of unknown function. Reduced transfer efficiency of plasmid from SalⅠ R-/M-to Sal Ⅰ R/M hosts argues that transfer of both the circular and linear forms of the plasmid involves double-stranded DNA. These results suggest that conjugal transfer occurs by a similar mechanism for SLP2-derived linear and circular plasmids, and cellular membrane/wall functions in the transfer process.