In a recent Nature Communications paper, Rice University professor James Chappell and collaborators introduced a novel way to improve plasmids—circular DNA molecules essential to molecular biology research since the 1970s. Instead of designing experiments around plasmids’ long‑standing limitations, the team redesigned part of the plasmid itself, specifically the origin of replication (ORI), which governs how plasmids duplicate inside cells.
“For decades, we’ve been designing experiments around two major limitations of plasmids: fixed copy numbers and incompatibility,” said Chappell. “While functional, such workarounds are clunky. We created a synthetic version of a part of the plasmid called the origin of replication that allows us to modify the plasmid instead of modifying the experiment.”
Typically, plasmids placed in bacterial cells use the cell’s internal machinery to replicate and produce proteins. Each plasmid generates fragments of a stop signal, called negative regulators, that attach to the ORI. When a sufficient number of regulators bind, plasmid replication ceases, maintaining a constant number of copies per cell. This copy number directly influences protein production—more plasmids yield more protein. Achieving reliable control over this process is vital for experiment design and outcomes.
Another persistent challenge is ORI incompatibility. There are roughly 27 classes of ORIs, and if two plasmids with similar ORIs coexist in one cell, their negative regulators mix, lowering protein production. This limitation typically prevents researchers from using several plasmids together.
To address these issues, Chappell’s team, in partnership with Matt Lakin at the University of New Mexico, engineered a synthetic ORI divided into two functional modules. One module dictates which stop signals are used, and the other sets how many are required to stop replication. This gives investigators precise control over copy number and compatibility within the same cell.
“Instead of using the natural stop signals, we used synthetically engineered RNA control elements,” said Baiyang Liu, first author of the study. “We have large libraries of unique RNA control elements that can be used in plasmids, meaning that we can potentially put large numbers of plasmids into a cell without incompatibility issues affecting plasmid replication.”
Testing showed that six plasmids with synthetic ORIs successfully replicated to their programmed copy numbers and expressed proteins as intended. The synthetic ORIs also responded dynamically to changes in cell conditions. As Chappell noted, the modular design “lets each researcher modify the plasmids to fit their experiments, simplifying their workflow and expanding experimental possibilities.”
Publication details: Liu, B., Seet, Z.R.D., Peng, X. et al. Engineering plasmids with synthetic origins of replication. Nat Commun 17, 2255 (2026). https://doi.org/10.1038/s41467-026-68907-1
“For decades, we’ve been designing experiments around two major limitations of plasmids: fixed copy numbers and incompatibility,” said Chappell. “While functional, such workarounds are clunky. We created a synthetic version of a part of the plasmid called the origin of replication that allows us to modify the plasmid instead of modifying the experiment.”
Typically, plasmids placed in bacterial cells use the cell’s internal machinery to replicate and produce proteins. Each plasmid generates fragments of a stop signal, called negative regulators, that attach to the ORI. When a sufficient number of regulators bind, plasmid replication ceases, maintaining a constant number of copies per cell. This copy number directly influences protein production—more plasmids yield more protein. Achieving reliable control over this process is vital for experiment design and outcomes.
Another persistent challenge is ORI incompatibility. There are roughly 27 classes of ORIs, and if two plasmids with similar ORIs coexist in one cell, their negative regulators mix, lowering protein production. This limitation typically prevents researchers from using several plasmids together.
To address these issues, Chappell’s team, in partnership with Matt Lakin at the University of New Mexico, engineered a synthetic ORI divided into two functional modules. One module dictates which stop signals are used, and the other sets how many are required to stop replication. This gives investigators precise control over copy number and compatibility within the same cell.
“Instead of using the natural stop signals, we used synthetically engineered RNA control elements,” said Baiyang Liu, first author of the study. “We have large libraries of unique RNA control elements that can be used in plasmids, meaning that we can potentially put large numbers of plasmids into a cell without incompatibility issues affecting plasmid replication.”
Testing showed that six plasmids with synthetic ORIs successfully replicated to their programmed copy numbers and expressed proteins as intended. The synthetic ORIs also responded dynamically to changes in cell conditions. As Chappell noted, the modular design “lets each researcher modify the plasmids to fit their experiments, simplifying their workflow and expanding experimental possibilities.”
Publication details: Liu, B., Seet, Z.R.D., Peng, X. et al. Engineering plasmids with synthetic origins of replication. Nat Commun 17, 2255 (2026). https://doi.org/10.1038/s41467-026-68907-1