ArticleNucleic acids research2017
Exponential propagation of large circular DNA by reconstitution of a chromosome-replication cycle.
Article in Nucleic acids research, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 37 papers.
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Who cites it
37 citing papers in PubMed.
- A self-replicating artificial module-genome that generates bacterial chromosome replication system in vitro.Nucleic acids research · 2026Article
- Transient Transfection of Rolling-Circle Amplified DNA in Biomanufacturing-Relevant Mammalian Cell Lines: A Comparison of Transfection Conditions for Optimal Protein Expression.Biotechnology and applied biochemistry · 2026Article
- Automated linear DNA assembly of A. thaliana's chloroplast and mitochondrial genome.Nucleic acids research · 2026Article
- Reconstitution of DNA fragments on HAC/MAC via the fragment-assembly system.Scientific reports · 2026Article
- Rapid immunization and antibody redesign platform discovers broadly neutralizing antibodies against non-immunized SARS-CoV-2 variant.Molecular therapy : the journal of the American Society of Gene Therapy · 2026Article
- Article
- Reconstituting alternative life using the test-bed of cell-free systems.Philosophical transactions of the Royal Society of London. Series B, Biological sciences · 2025Review
- Synthesizing supercoiled circular DNA molecules in vitro.Nucleic acids research · 2025Article
- Simultaneous in vitro expression of minimal 21 transfer RNAs by tRNA array method.Nature communications · 2025Article
- Adaptive evolution of oriC through in vitro propagation of a mini-chromosome in RCR.Nucleic acids research · 2025Article
- Sustainable regeneration of 20 aminoacyl-tRNA synthetases in a reconstituted system toward self-synthesizing artificial systems.Science advances · 2025Article
- Self-growing protocell models in aqueous two-phase system induced by internal DNA replication reaction.Nature communications · 2025Article
- Real-time analysis of initiation regulation systems during the progression of the reconstituted chromosomal replication cycle.Scientific reports · 2025Article
- Autonomous ribosome biogenesis in vitro.Nature communications · 2025Article
- Cell-Free Gene Expression: Methods and Applications.Chemical reviews · 2025Review
- Efficient variant phasing utilizing a replication cycle reaction system.Genetics in medicine open · 2025Article
- Establishing a biochemical understanding of the initiation of chromosome replication in bacteria.Proceedings of the National Academy of Sciences of the United States of America · 2024Article
- Synthetic Genomics: Repurposing Biological Systems for Applications in Engineering Biology.ACS synthetic biology · 2024Review
- Building Synthetic Cells─From the Technology Infrastructure to Cellular Entities.ACS synthetic biology · 2024Review
- In vitro one-pot construction of influenza viral genomes for virus particle synthesis based on reverse genetics system.PloS one · 2024Article
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4 authors.
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No grant is acknowledged in the PubMed record.
Abstract
Propagation of genetic information is a fundamental property of living organisms. Escherichia coli has a 4.6 Mb circular chromosome with a replication origin, oriC. While the oriC replication has been reconstituted in vitro more than 30 years ago, continuous repetition of the replication cycle has not yet been achieved. Here, we reconstituted the entire replication cycle with 14 purified enzymes (25 polypeptides) that catalyze initiation at oriC, bidirectional fork progression, Okazaki-fragment maturation and decatenation of the replicated circular products. Because decatenation provides covalently closed supercoiled monomers that are competent for the next round of replication initiation, the replication cycle repeats autonomously and continuously in an isothermal condition. This replication-cycle reaction (RCR) propagates ∼10 kb circular DNA exponentially as intact covalently closed molecules, even from a single DNA molecule, with a doubling time of ∼8 min and extremely high fidelity. Very large DNA up to 0.2 Mb is successfully propagated within 3 h. We further demonstrate a cell-free cloning in which RCR selectively propagates circular molecules constructed by a multi-fragment assembly reaction. Our results define the minimum element necessary for the repetition of the chromosome-replication cycle, and also provide a powerful in vitro tool to generate large circular DNA molecules without relying on conventional biological cloning.
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