ArticleScientific reports2018
Isolation and characterization of novel mutations in the pSC101 origin that increase copy number.
Article in Scientific reports, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 25 papers.
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25 citing papers in PubMed.
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- Binary vector copy number engineering improves Agrobacterium-mediated transformation.Nature biotechnology · 2025Article
- Deep learning guided programmable design of Escherichia coli core promoters from sequence architecture to strength control.Nucleic acids research · 2025Article
- How many plasmids can bacteria carry? A synthetic biology perspective.Open biology · 2025Review
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- Probing the orthogonality and robustness of the mammalian RNA-binding protein Musashi-1 in Escherichia coli.Journal of biological engineering · 2024Article
- Engineering plasmid copy number heterogeneity for dynamic microbial adaptation.Nature microbiology · 2024Article
- Live-cell imaging reveals the trade-off between target search flexibility and efficiency for Cas9 and Cas12a.Nucleic acids research · 2024Article
- Toxin/antitoxin systems induce persistence and work in concert with restriction/modification systems to inhibit phage.Microbiology spectrum · 2024Article
- CRISPRi-mediated tunable control of gene expression level with engineered single-guide RNA in Escherichia coli.Nucleic acids research · 2023Article
- Inducible plasmid copy number control for synthetic biology in commonly used E. coli strains.Nature communications · 2022Article
- Inducible directed evolution of complex phenotypes in bacteria.Nucleic acids research · 2022Article
- Single cell variability of CRISPR-Cas interference and adaptation.Molecular systems biology · 2022Article
- Strategies for Improving Small-Molecule Biosensors in Bacteria.Biosensors · 2022Review
- Increased copy number couples the evolution of plasmid horizontal transmission and plasmid-encoded antibiotic resistance.Proceedings of the National Academy of Sciences of the United States of America · 2021Article
- Beyond horizontal gene transfer: the role of plasmids in bacterial evolution.Nature reviews. Microbiology · 2021Review
- Article
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9 authors.
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Abstract
pSC101 is a narrow host range, low-copy plasmid commonly used for genetically manipulating Escherichia coli. As a byproduct of a genetic screen for a more sensitive lactam biosensor, we identified multiple novel mutations that increase the copy number of plasmids with the pSC101 origin. All mutations identified in this study occurred on plasmids which also contained at least one mutation localized to the RepA protein encoded within the origin. Homology modelling predicts that many of these mutations occur within the dimerization interface of RepA. Mutant RepA resulted in plasmid copy numbers between ~31 and ~113 copies/cell, relative to ~5 copies/cell in wild-type pSC101 plasmids. Combining the mutations that were predicted to disrupt multiple contacts on the dimerization interface resulted in copy numbers of ~500 copies/cell, while also attenuating growth in host strains. Fluorescent protein production expressed from an arabinose-inducible promoter on mutant origin derived plasmids did correlate with copy number. Plasmids harboring RepA with one of two mutations, E83K and N99D, resulted in fluorescent protein production similar to that from p15a- (~20 copies/cell) and ColE1- (~31 copies/cell) based plasmids, respectively. The mutant copy number variants retained compatibility with p15a, pBBR, and ColE1 origins of replication. These pSC101 variants may be useful in future metabolic engineering efforts that require medium or high-copy vectors compatible with p15a- and ColE1-based plasmids.
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