ArticleMethods in molecular biology (Clifton, N.J.)2012
Quantifying plasmid copy number to investigate plasmid dosage effects associated with directed protein evolution.
Article in Methods in molecular biology (Clifton, N.J.), 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 10 papers.
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Who cites it
10 citing papers in PubMed.
- Plasmid mutation rates scale with copy number.Proceedings of the National Academy of Sciences of the United States of America · 2026Article
- The plasmid-host fitness landscape: a new paradigm for predicting the fate of mobile resistance.Applied and environmental microbiology · 2026Review
- How many plasmids can bacteria carry? A synthetic biology perspective.Open biology · 2025Review
- The polyadenylase PAPI is required for virulence plasmid maintenance in pathogenic bacteria.PLoS pathogens · 2025Article
- Deletion of pcnB affects antibiotic susceptibility in resistant Escherichia coli by reducing copy number of ColE1-family plasmids.Scientific reports · 2025Article
- Segregational instability of multicopy plasmids: A population genetics approach.Ecology and evolution · 2022Article
- Beyond horizontal gene transfer: the role of plasmids in bacterial evolution.Nature reviews. Microbiology · 2021Review
- Genetic control of ColE1 plasmid stability that is independent of plasmid copy number regulation.Current genetics · 2019Article
- Engineering posttranslational proofreading to discriminate nonstandard amino acids.Proceedings of the National Academy of Sciences of the United States of America · 2018Article
- Chromatic Bacteria - A Broad Host-Range Plasmid and Chromosomal Insertion Toolbox for Fluorescent Protein Expression in Bacteria.Frontiers in microbiology · 2018Article
Corrections and comments
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Authors and funding
4 authors.
Funding
Abstract
Our laboratory specializes in directed protein evolution, i.e., evolution of proteins under defined selective pressures in the laboratory. Our target genes are encoded in ColE1 plasmids to facilitate the generation of libraries in vivo. We have observed that when random mutations are not restricted to the coding sequence of the target genes, directed evolution results in a strong positive selection of plasmid origin of replication (ori) mutations. Surprisingly, this is true even during evolution of new biochemical activities, when the activity that is being selected was not originally present. The selected plasmid ori mutations are diverse and produce a range of plasmid copy numbers, suggesting a complex interplay between ori and coding mutations rather than a simple enhancement of level of expression of the target gene. Thus, plasmid dosage may contribute significantly to evolution by fine-tuning levels of activity. Here, we present examples illustrating these observations as well as our methods for efficient quantification of plasmid copy number.
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