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Laboratory evolution and polyploid SCRaMbLE reveal genomic plasticity to synthetic chromosome defects and rearrangements

View ORCID ProfileThomas C. Williams, Heinrich Kroukamp, Xin Xu, Elizabeth L. I. Wightman, Briardo Llorente, Anthony R. Borneman, Alexander C. Carpenter, Niel Van Wyk, Monica I. Espinosa, Elizabeth L. Daniel, Roy S. K. Walker, Yizhi Cai, Helena K. M. Nevalainen, Natalie C. Curach, View ORCID ProfileIra W. Deveson, Timothy R. Mercer, Daniel L. Johnson, Leslie A. Mitchell, View ORCID ProfileJoel S. Bader, View ORCID ProfileGiovanni Stracquadanio, View ORCID ProfileJef D. Boeke, Hugh D. Goold, Isak S. Pretorius, Ian T. Paulsen
doi: https://doi.org/10.1101/2022.07.22.501046
Thomas C. Williams
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
2CSIRO Synthetic Biology Future Science Platform, Canberra, ACT 2601, Australia
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  • ORCID record for Thomas C. Williams
  • For correspondence: tom.williams@mq.edu.au sakkie.pretorius@mq.edu.au ian.paulsen@mq.edu.au
Heinrich Kroukamp
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
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Xin Xu
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
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Elizabeth L. I. Wightman
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
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Briardo Llorente
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
2CSIRO Synthetic Biology Future Science Platform, Canberra, ACT 2601, Australia
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Anthony R. Borneman
3The Australian Wine Research Institute, Adelaide, SA 5064, Australia
4School of Agriculture, Food & Wine, Faculty of Sciences, University of Adelaide, Adelaide, SA 5005, Australia
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Alexander C. Carpenter
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
2CSIRO Synthetic Biology Future Science Platform, Canberra, ACT 2601, Australia
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Niel Van Wyk
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
5Department of Microbiology and Biochemistry, Hochschule Geisenheim University, Geisenheim, Germany
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Monica I. Espinosa
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
2CSIRO Synthetic Biology Future Science Platform, Canberra, ACT 2601, Australia
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Elizabeth L. Daniel
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
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Roy S. K. Walker
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
6School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, Scotland, UK.
7School of Engineering, Institute for Bioengineering, The University of Edinburgh, Edinburgh, EH9 3BF, Scotland
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Yizhi Cai
6School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, Scotland, UK.
8SYNBIOCHEM, Manchester Institute of Biotechnology and School of Chemistry, University of Manchester, Manchester M13 9PL, UK.
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Helena K. M. Nevalainen
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
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Natalie C. Curach
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
9Bioplatforms Australia, Research Park Drive, Macquarie University, Macquarie Park, NSW 2109, Australia.
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Ira W. Deveson
10St Vincent’s Clinical School, University of New South Wales, Sydney, NSW 2010, Australia
11The Kinghorn Centre for Clinical Genomics, Garvan Institute of Medical Research, Sydney, NSW 2010, Australia
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Timothy R. Mercer
10St Vincent’s Clinical School, University of New South Wales, Sydney, NSW 2010, Australia
11The Kinghorn Centre for Clinical Genomics, Garvan Institute of Medical Research, Sydney, NSW 2010, Australia
12Australian Institute for Bioengineering and Nanotechnology, University of Queensland, Brisbane, QLD, Australia
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Daniel L. Johnson
3The Australian Wine Research Institute, Adelaide, SA 5064, Australia
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Leslie A. Mitchell
13Institute for Systems Genetics, NYU Langone Health, New York, NY 10016
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Joel S. Bader
14Department of Biomedical Engineering, Johns Hopkins University, 3400 North Charles Street, Baltimore, MD 21218
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Giovanni Stracquadanio
15Institute of Quantitative Biology, Biochemistry, and Biotechnology, SynthSys, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK
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Jef D. Boeke
13Institute for Systems Genetics, NYU Langone Health, New York, NY 10016
16Department of Biochemistry and Molecular Pharmacology, NYU Langone Health, New York, NY 10016
17Department of Biomedical Engineering, NYU Tandon School of Engineering, Brooklyn 11201, NY
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Hugh D. Goold
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
18New South Wales Department of Primary Industries, Orange, NSW 2800, Australia
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Isak S. Pretorius
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
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  • For correspondence: tom.williams@mq.edu.au sakkie.pretorius@mq.edu.au ian.paulsen@mq.edu.au
Ian T. Paulsen
1School of Natural Sciences, and ARC Centre of Excellence in Synthetic Biology, Macquarie University, Sydney, Australia
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  • For correspondence: tom.williams@mq.edu.au sakkie.pretorius@mq.edu.au ian.paulsen@mq.edu.au
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Summary

We have designed, constructed, and debugged a synthetic 753,096 bp version of Saccharomyces cerevisiae chromosome XIV as part of the international Sc2.0 project. We showed that certain synthetic loxPsym recombination sites can interfere with mitochondrial protein localization, that the deletion of one intron (NOG2) reduced fitness, and that a reassigned stop codon can lead to a growth defect. In parallel to these rational debugging modifications, we used Adaptive Laboratory Evolution to generate a general growth defect suppressor rearrangement in the form of increased TAR1 copy number. We also extended the utility of the Synthetic Chromosome Recombination and Modification by LoxP-mediated Evolution (SCRaMbLE) system by engineering synthetic-wild-type tetraploid hybrid strains that buffer against essential gene loss. The presence of wild-type chromosomes in the hybrid tetraploids increased post-SCRaMbLE viability and heterologous DNA integration, highlighting the plasticity of the S. cerevisiae genome in the presence of rational and non-rational modifications.

Competing Interest Statement

TCW and ACC are founders and shareholders of Number 8 Bio Pty Ltd. JDB is a Founder and Director of CDI Labs, Inc., a Founder of and consultant to Neochromosome, Inc, a Founder, SAB member of and consultant to ReOpen Diagnostics, LLC and serves or served on the Scientific Advisory Board of the following: Sangamo, Inc., Modern Meadow, Inc., Rome Therapeutics, Inc., Sample6, Inc., Tessera Therapeutics, Inc. and the Wyss Institute. JSB is a Founder of Neochromosome, Inc., and a consultant to Opentrons Labworks, Inc. LAM is a Founder of Neochromosome, Inc., and an employee of Opentrons Labworks, Inc.

Footnotes

  • ↵‡ Present affiliations

  • 'Elizabteh' was corrected to 'Elizabeth' in the author list.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY 4.0 International license.
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Laboratory evolution and polyploid SCRaMbLE reveal genomic plasticity to synthetic chromosome defects and rearrangements
Thomas C. Williams, Heinrich Kroukamp, Xin Xu, Elizabeth L. I. Wightman, Briardo Llorente, Anthony R. Borneman, Alexander C. Carpenter, Niel Van Wyk, Monica I. Espinosa, Elizabeth L. Daniel, Roy S. K. Walker, Yizhi Cai, Helena K. M. Nevalainen, Natalie C. Curach, Ira W. Deveson, Timothy R. Mercer, Daniel L. Johnson, Leslie A. Mitchell, Joel S. Bader, Giovanni Stracquadanio, Jef D. Boeke, Hugh D. Goold, Isak S. Pretorius, Ian T. Paulsen
bioRxiv 2022.07.22.501046; doi: https://doi.org/10.1101/2022.07.22.501046
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Laboratory evolution and polyploid SCRaMbLE reveal genomic plasticity to synthetic chromosome defects and rearrangements
Thomas C. Williams, Heinrich Kroukamp, Xin Xu, Elizabeth L. I. Wightman, Briardo Llorente, Anthony R. Borneman, Alexander C. Carpenter, Niel Van Wyk, Monica I. Espinosa, Elizabeth L. Daniel, Roy S. K. Walker, Yizhi Cai, Helena K. M. Nevalainen, Natalie C. Curach, Ira W. Deveson, Timothy R. Mercer, Daniel L. Johnson, Leslie A. Mitchell, Joel S. Bader, Giovanni Stracquadanio, Jef D. Boeke, Hugh D. Goold, Isak S. Pretorius, Ian T. Paulsen
bioRxiv 2022.07.22.501046; doi: https://doi.org/10.1101/2022.07.22.501046

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