Identifying and characterizing SCRaMbLEd synthetic yeast using ReSCuES
2018
SCRaMbLEis a novel system implemented in the synthetic yeast genome, enabling massive
chromosome rearrangementsto produce strains with a large genotypic diversity upon induction. Here we describe a reporter of
SCRaMbLEdcells using efficient selection, termed ReSCuES, based on a loxP-mediated switch of two auxotrophic markers. We show that all randomly isolated clones contained rearrangements within the synthetic chromosome, demonstrating high efficiency of selection. Using ReSCuES, we illustrate the ability of
SCRaMbLEto generate strains with increased tolerance to several stress factors, such as ethanol, heat and acetic acid. Furthermore, by analyzing the tolerant strains, we are able to identify ACE2, a transcription factor required for septum destruction after
cytokinesis, as a negative regulator of ethanol tolerance. Collectively, this work not only establishes a generic platform to rapidly identify strains of interest by
SCRaMbLE, but also provides methods to dissect the underlying mechanisms of resistance.
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