RT Journal Article
SR Electronic
T1 Preventing plasmid multimer formation in commonly used synthetic biology plasmids
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2024.07.23.604805
DO 10.1101/2024.07.23.604805
A1 Vaisbourd, Elizabeth
A1 Bren, Anat
A1 Alon, Uri
A1 Glass, David S.
YR 2024
UL http://biorxiv.org/content/early/2024/07/24/2024.07.23.604805.abstract
AB Plasmids are an essential tool for basic research and biotechnology applications. To optimize plasmid-based circuits, it is crucial to control plasmid integrity, including the formation of plasmid multimers. Multimers are tandem repeats of entire plasmids formed during replication by failed dimer resolution. Multimers can affect the behavior of synthetic circuits, especially ones that include DNA-editing enzymes. However, occurrence of multimers is not commonly assayed. Here we survey four commonly used plasmid backbones for occurrence of multimers in cloning (JM109) and wild-type (MG1655) strains. We find that multimers occur appreciably only in MG1655, with the fraction of plasmids existing as multimers increasing with both plasmid copy number and culture passaging. In contrast, introduction of multimers into JM109 can produce strains containing only multimers. We present an MG1655 ΔrecA single-locus knockout that avoids multimer production. These results can aid synthetic biologists in improving design and reliability of plasmid-based circuits.Competing Interest StatementThe authors have declared no competing interest.