Characterization of endogenous plasmids from Lactobacillus salivarius UCC118.
The genome of Lactobacillus salivarius UCC118 comprises a 1.83-Mb chromosome, a 242-kb megaplasmid (pMP118), and two smaller plasmids of 20 kb (pSF118-20) and 44 kb (pSF118-44). Annotation and bioinformatic analyses suggest that both of the smaller plasmids replicate by a theta replication mechanism. Furthermore, it appears that they are transmissible, although neither possesses a complete set of conjugation genes. Plasmid pSF118-20 encodes a toxin-antitoxin system composed of pemI and pemK homologs, and this plasmid could be cured when PemI was produced in trans. The minimal replicon of pSF118-20 was determined by deletion analysis. Shuttle vector derivatives of pSF118-20 were generated that included the replication region (pLS203) and the replication region plus mobilization genes (pLS208). The plasmid pLS203 was stably maintained without selection in Lactobacillus plantarum, Lactobacillus fermentum, and the pSF118-20-cured derivative strain of L. salivarius UCC118 (strain LS201). Cloning in pLS203 of genes encoding luciferase and green fluorescent protein, and expression from a constitutive L. salivarius promoter, demonstrated the utility of this vector for the expression of heterologous genes in Lactobacillus. This study thus expands the knowledge base and vector repertoire of probiotic lactobacilli.
Fang
Fang
F
Department of Microbiology, University College Cork, Cork, Ireland.
Flynn
Sarah
S
Li
Yin
Y
Claesson
Marcus J
MJ
van Pijkeren
Jan-Peter
JP
Collins
J Kevin
JK
van Sinderen
Douwe
D
O'Toole
Paul W
PW
D002874:Chromosome Mapping; D003001:Cloning, Molecular; D003227:Conjugation, Genetic; D004261:DNA Replication / Q000235:genetics; D017353:Gene Deletion; D023061:Gene Order; D005798:Genes, Bacterial; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D007778:Lactobacillus / Q000235:genetics*; D008156:Luciferases / Q000235:genetics; D020161:Physical Chromosome Mapping; D010957:Plasmids / Q000302:isolation & purification*; D012093:Replicon
D049452:Green Fluorescent Proteins; D008156:Luciferases