Characterization of a cryptic plasmid from Bacillus sphaericus strain LP1-G
Enying Wu1, Liang Jun, Yongming Yuan
1Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China.
Plasmid
|January 16, 2007
Summary
The Bacillus sphaericus plasmid pLG was characterized as an 11,066bp molecule belonging to the rolling-circle replication (RCR) pC194-family. Its replication mechanism and genetics were elucidated, revealing a minimal replicon of 1.6kb.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Bacillus sphaericus harbors cryptic plasmids that require characterization for understanding bacterial genetics.
- Rolling-circle replication (RCR) is a common plasmid replication mechanism in bacteria.
Purpose of the Study:
- To sequence and characterize the cryptic plasmid pLG from Bacillus sphaericus strain LP1-G.
- To elucidate the replication mechanism and genetic elements of the pLG plasmid.
Main Methods:
- Plasmid sequencing and bioinformatic analysis to identify Open Reading Frames (ORFs).
- Sequence analysis to identify replication origins (dso and sso).
- Mung bean nuclease analysis and Southern hybridization to detect single-stranded DNA (ssDNA) intermediates.
- Subcloning and transformation experiments to determine the minimal replicon.
Main Results:
- The pLG plasmid is an 11,066bp circular molecule with a G+C content of 37%, encoding 23 putative ORFs.
- ORF 21 shows homology to Rep proteins of the RCR pC194-family, with identified pC194-type dso and sso sequences.
- ssDNA intermediates were confirmed, indicating RCR replication. A unique replication mechanism is suggested by ssDNA accumulation patterns.
- The minimal replicon, consisting of the rep gene and dso, is approximately 1.6kb. The plasmid copy number is about 58 per cell.
Conclusions:
- pLG belongs to the RCR pC194-family, utilizing a double-strand origin (dso) and single-strand origin (sso) for replication.
- The study provides a foundation for understanding the replication and genetics of this Bacillus sphaericus plasmid.
- Potential unique regulatory mechanisms governing pLG replication warrant further investigation.
Related Concept Videos
01:28Plasmids
Plasmids are extrachromosomal DNA molecules found in bacteria, archaea, and some eukaryotic microbes like yeast. These small, circular DNA structures typically contain fewer than 30 genes, although some may exist linearly. Plasmids vary in their number within a cell, known as copy number. Single-copy plasmids are present in one copy per cell and multi-copy plasmids are present in multiple copies, reaching over 100 copies per cell.Plasmids usually replicate independently of the chromosomal DNA...
01:26DNA Bacteriophages
Bacteriophages, or phages, are viruses that specifically infect bacteria, utilizing their genetic material to hijack host cellular machinery for replication. DNA bacteriophages employ single-stranded DNA (ssDNA) or double-stranded DNA (dsDNA) genomes. These phages exhibit diverse replication strategies and host interactions, influencing their ecological roles and applications in biotechnology and medicine.ssDNA BacteriophagesssDNA phages, with their small genomes, utilize unique strategies to...
01:16Viral Replication: Lysogenic Cycle
The lysogenic cycle is a crucial viral replication strategy that allows bacteriophages to persist within host cells without immediately destroying them. This process is primarily observed in temperate phages, such as bacteriophage lambda (λ), which infects Escherichia coli. The cycle allows the viral genome to persist across bacterial generations while keeping host cells viable.Integration of the Viral GenomeUpon infection, bacteriophage lambda attaches to the bacterial surface and injects its...
01:29Coordination of Gene Expression Processes in Bacteria
The DNA replication, transcription, and translation processes are intricately coupled in bacteria, allowing efficient gene expression and rapid protein synthesis. While this physical and functional coordination is advantageous, it introduces challenges that bacteria overcome through specific regulatory mechanisms.Coupling of Replication, Transcription, and TranslationThe coupling of replication, transcription, and translation is a hallmark of bacterial gene expression. As the replisome unwinds...