Characterization of a novel rolling-circle replication plasmid pYSI8 from Lactobacillus sakei YSI8.
A plasmid from Lactobacillus sakei YSI8, designated as pYSI8, was sequenced and characterized. It consisted of a 4973bp circular molecule with a G+C content of 35.6%. The plasmid pYSI8 was predicted to contain five putative ORFs, in which ORF1 shared 79% and 76% identity with Rep proteins of pLH2 and pLC2, members of rolling-circle replication (RCR) pMV158 family. Detection of single-stranded DNA (ssDNA) intermediates by Southern hybridization and mung bean nuclease treatment confirmed that pYSI8 replicated via the RCR mechanism. Accumulation of ssDNA in rifampicin-treated strains implied that the host-encoded RNA polymerase was involved in the conversion of ssDNA to double-stranded DNA (dsDNA). Furthermore, the copy number of pYSI8 was estimated to be 41.9+/-0.5 in each cell by real-time polymerase chain reaction.
Zhai
Zhengyuan
Z
College of Food Science and Nutritional Engineering, China Agricultural University, 17 Qing Hua East Road, Hai Dian District, Beijing 100083, China.
Hao
Yanling
Y
Yin
Sheng
S
Luan
Chunguang
C
Zhang
Liebing
L
Zhao
Lu
L
Chen
Dong
D
Wang
Ou
O
Luo
Yunbo
Y
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry; D001483:Base Sequence; D004261:DNA Replication; D004277:DNA, Single-Stranded / Q000235:genetics; D018628:Gene Dosage; D007778:Lactobacillus / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics*; D018741:Replication Origin / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded