Synthesis, debugging, and effects of synthetic chromosome consolidation: synVI and beyond

Abstract

We describe design, rapid assembly, and characterization of synthetic yeast Sc2.0 chromosome VI (synVI). A mitochondrial defect in the synVI strain mapped to synonymous coding changes within <i>PRE4</i> (<i>YFR050C</i>), encoding an essential proteasome subunit; Sc2.0 coding changes reduced Pre4 protein accumulation by half. Completing Sc2.0 specifies consolidation of 16 synthetic chromosomes into a single strain. We investigated phenotypic, transcriptional, and proteomewide consequences of Sc2.0 chromosome consolidation in poly-synthetic strains. Another "bug" was discovered through proteomic analysis, associated with alteration of the <i>HIS2</i> transcription start due to transfer RNA deletion and loxPsym site insertion. Despite extensive genetic alterations across 6% of the genome, no major global changes were detected in the poly-synthetic strain "omics" analyses. This work sets the stage for completion of a designer, synthetic eukaryotic genome.