Synthetic yeast chromosome XI design provides a testbed for the study of extrachromosomal circular DNA dynamics

Blount, Benjamin A; Lu, Xinyu; Driessen, Maureen R M; Jovicevic, Dejana; Sanchez, Mateo I; Ciurkot, Klaudia; Zhao, Yu; Lauer, Stephanie; McKiernan, Robert M; Gowers, Glen-Oliver F; Sweeney, Fiachra; Fanfani, Viola; Lobzaev, Evgenii; Palacios-Flores, Kim; Walker, Roy S K; Hesketh, Andy; Cai, Jitong; Oliver, Stephen G; Cai, Yizhi; Stracquadanio, Giovanni; Mitchell, Leslie A; Bader, Joel S; Boeke, Jef D; Ellis, Tom

Abstract

We describe construction of the synthetic yeast chromosome XI (synXI) and reveal the effects of redesign at non-coding DNA elements. The 660-kb synthetic yeast genome project (Sc2.0) chromosome was assembled from synthesized DNA fragments before CRISPR-based methods were used in a process of bug discovery, redesign, and chromosome repair, including precise compaction of 200 kb of repeat sequence. Repaired defects were related to poor centromere function and mitochondrial health and were associated with modifications to non-coding regions. As part of the Sc2.0 design, loxPsym sequences for Cre-mediated recombination are inserted between most genes. Using the GAP1 locus from chromosome XI, we show that these sites can facilitate induced extrachromosomal circular DNA (eccDNA) formation, allowing direct study of the effects and propagation of these important molecules. Construction and characterization of synXI contributes to our understanding of non-coding DNA elements, provides a useful tool for eccDNA study, and will inform future synthetic genome design.

Citation

Blount, B. A., Lu, X., Driessen, M. R. M., Jovicevic, D., Sanchez, M. I., Ciurkot, K., Zhao, Y., Lauer, S., McKiernan, R. M., Gowers, G.-O. F., Sweeney, F., Fanfani, V., Lobzaev, E., Palacios-Flores, K., Walker, R. S. K., Hesketh, A., Cai, J., Oliver, S. G., Cai, Y., Stracquadanio, G., …Ellis, T. (2023). Synthetic yeast chromosome XI design provides a testbed for the study of extrachromosomal circular DNA dynamics. Cell Genomics, 3(11), Article 100418. https://doi.org/10.1016/j.xgen.2023.100418

• Journal Article Type: Article
• Acceptance Date: Sep 8, 2023
• Online Publication Date: Nov 9, 2023
• Publication Date: Nov 9, 2023
• Deposit Date: Nov 10, 2023
• Publicly Available Date: Nov 10, 2023
• Journal: Cell Genomics
• Electronic ISSN: 2666-979X
• Publisher: Cell Press
• Peer Reviewed: Peer Reviewed
• Volume: 3
• Issue: 11
• Article Number: 100418
• DOI: https://doi.org/10.1016/j.xgen.2023.100418
• Keywords: Synthetic biology; synthetic genomics; extrachromosomal circular DNA; eccDNA; Sc2.0; Yeast 2.0; genome editing; stress adaptation
• Public URL: https://nottingham-repository.worktribe.com/output/27086039
• Publisher URL: https://www.sciencedirect.com/science/article/pii/S2666979X23002446?via%3Dihub
• Additional Information: This article is maintained by: Elsevier; Article Title: Synthetic yeast chromosome XI design provides a testbed for the study of extrachromosomal circular DNA dynamics; Journal Title: Cell Genomics; CrossRef DOI link to publisher maintained version: https://doi.org/10.1016/j.xgen.2023.100418; Content Type: article; Copyright: © 2023 The Author(s).