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; Niël van Wyk; Monica I. Espinosa; Elizabeth L. Daniel; Roy S. K. Walker; Yizhi Cai; Helena K. M. Nevalainen; Natalie C. Curach; Ira W. Deveson; Tim R. Mercer; Daniel L. Johnson; LESLIE ANN MITCHELL; Joel S. Bader; Giovanni Stracquadanio; Jef D. Boeke; Hugh Douglas Goold; Isak S. Pretorius; Ian T. Paulsen

doi:10.1101/2022.07.22.501046

Synthetic BiologybioRxivJul 22, 2022

Preprint

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

Thomas C. Williams iD, Heinrich Kroukamp iD, Xin Xu iD, Elizabeth L. I. Wightman iD, Briardo Llorente iD, Anthony R. Borneman iD, Alexander C. Carpenter iD, Niël van Wyk iD, Monica I. Espinosa iD, Elizabeth L. Daniel, Roy S. K. Walker iD, Yizhi Cai iD, Helena K. M. Nevalainen iD, Natalie C. Curach iD, Ira W. Deveson iD, Tim R. Mercer iD, Daniel L. Johnson iD, LESLIE ANN MITCHELL iD, Joel S. Bader iD, Giovanni Stracquadanio iD, Jef D. Boeke iD, Hugh Douglas Goold iD, Isak S. Pretorius iD, Ian T. Paulsen iD

Abstract

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.

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