Mind your marker: A genome-wide analysis of the effect of common auxotrophic markers on complex traits in yeast

Yeast are extensively used as a model organism for studying human diseases and screening for novel therapeutics due to their highly conserved genome and cell biology. Indeed, studies in yeast models have led to several breakthroughs in understanding aging, neurodegeneration, cancer, metabolic disorders, and others. While many cellular processes are conserved among yeast and higher organisms, including humans, most common laboratory strains include mutations in genes encoding for enzymes in the biosynthetic pathways of several core metabolites, such as the amino acids leucine, histidine, methionine, tryptophan, and lysine, and the nucleobases uracil and adenine. These genetic perturbations, known as auxotrophic markers, are powerful and fundamental tools in yeast genetic engineering, as they allow to readily select positive transformants on synthetic defined media lacking the specific metabolite.

Previous studies have reported that when uncompensated, autotrophic markers cause significant phenotypic bias as compared to prototrophic strains and have different combinatorial influences on the metabolic network. Evidently, the metabolic background affects up to 85% of the coding genome, acting at the transcriptomic, proteomic, and metabolomic levels altogether. We have genomically edited the popular laboratory yeast strain, BY4741, to derive an endogenous strain collection comprising of all possible combinations of the four common auxotrophic markers leu2∆, ura3∆, his3∆, met15∆, that are involved in the synthesis of leucine, uracil, histidine, and methionine, respectively. As a proof of concept for the broad consequences of the metabolic background, we tested the effects of auxotrophic markers on yeast fitness at various environmental conditions.