Characterization of the cryptic plasmid pTXW from Lactobacillus paracasei TXW.
A cryptic plasmid from Lactobacillus paracasei TXW isolated from koumiss, designated as pTXW, was sequenced and characterized. It is 3178 bp in length with a G+C content of 42.9%. The plasmid pTXW was predicted to encode 4 putative ORFs. RepB shared high homology with initiator proteins of plasmids from the rolling circle replication (RCR) pMV158 family. copG was predicted to encode a transcriptional repressor, which was involved in plasmid copy number control together with a putative antisense RNA. Mob putatively acts as a relaxase and belongs to the MOB(V) relaxase family. Sequence analysis revealed a double strand origin (dso) located upstream the repB gene, which contained the conserved nick sequence of the pMV158 family. A putative single strand origin (sso-like) consisting of successive inverted repeats was also detected. Mung bean nuclease analysis and Southern hybridization confirmed the presence of single-stranded DNA (ssDNA) intermediates, suggesting that pTXW replicates via the RCR mechanism. Furthermore, the relative copy number of pTXW was estimated to be 73.5±1.2 copies in each cell by real-time PCR. The new RCR plasmid would be valuable to construct vectors for application in food industry.
Zhang
Hongxing
H
Department of Food Science, Beijing University of Agriculture, 7 Bei Nong Road, ChangPing District, Beijing 102206, China.
Hao
Yanling
Y
Zhang
Dapeng
D
Luo
Yunbo
Y
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry; D001483:Base Sequence; D015139:Blotting, Southern; D004277:DNA, Single-Stranded / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D018628:Gene Dosage / Q000235:genetics; D007778:Lactobacillus / Q000235:genetics*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D016366:Open Reading Frames / Q000235:genetics; D020161:Physical Chromosome Mapping; D010957:Plasmids / Q000235:genetics* / Q000302:isolation & purification*; D012015:Reference Standards; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded