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• W2017677183 abstract "ABSTRACT Xylella fastidiosa strain riv11 harbors a 25-kbp plasmid (pXF-RIV11) belonging to the IncP-1 incompatibility group. Replication and stability factors of pXF-RIV11 were identified and used to construct plasmids able to replicate in X. fastidiosa and Escherichia coli . Replication in X. fastidiosa required a 1.4-kbp region from pXF-RIV11 containing a replication initiation gene ( trfA ) and the adjacent origin of DNA replication ( oriV ). Constructs containing trfA and oriV from pVEIS01, a related IncP-1 plasmid of the earthworm symbiont Verminephrobacter eiseniae , also were competent for replication in X. fastidiosa . Constructs derived from pXF-RIV11 but not pVEIS01 replicated in Agrobacterium tumefaciens , Xanthomonas campestris , and Pseudomonas syringae . Although plasmids bearing replication elements from pXF-RIV11 or pVEIS01 could be maintained in X. fastidiosa under antibiotic selection, removal of selection resulted in plasmid extinction after 3 weekly passages. Addition of a toxin-antitoxin addiction system ( pemI / pemK ) from pXF-RIV11 improved plasmid stability such that >80 to 90% of X. fastidiosa cells retained plasmid after 5 weekly passages in the absence of antibiotic selection. Expression of PemK in E. coli was toxic for cell growth, but toxicity was nullified by coexpression of PemI antitoxin. Deletion of N-terminal sequences of PemK containing the conserved motif RGD abolished toxicity. In vitro assays revealed a direct interaction of PemI with PemK, suggesting that antitoxin activity of PemI is mediated by toxin sequestration. IncP-1 plasmid replication and stability factors were added to an E. coli cloning vector to constitute a stable 6.0-kbp shuttle vector (pXF20-PEMIK) suitable for use in X. fastidiosa ." @default.
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• W2017677183 date "2010-12-01" @default.
• W2017677183 modified "2023-09-26" @default.
• W2017677183 title "Functional Characterization of Replication and Stability Factors of an Incompatibility Group P-1 Plasmid from Xylella fastidiosa" @default.
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• W2017677183 doi "https://doi.org/10.1128/aem.01921-10" @default.
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