Genome-scale analysis of interactions between genetic perturbations and natural variation.

Interactions between genetic perturbations and segregating loci can cause perturbations to show different phenotypic effects across genetically distinct individuals. To study these interactions on a genome scale in many individuals, we used combinatorial DNA barcode sequencing to measure the fitness effects of 8046 CRISPRi perturbations targeting 1721 distinct genes in 169 yeast cross progeny (or segregants). We identified 460 genes whose perturbation has different effects across segregants. Several factors caused perturbations to show variable effects, including baseline segregant fitness, the mean effect of a perturbation across segregants, and interacting loci. We mapped 234 interacting loci and found four hub loci that interact with many different perturbations. Perturbations that interact with a given hub exhibit similar epistatic relationships with the hub and show enrichment for cellular processes that may mediate these interactions. These results suggest that an individual's response to perturbations is shaped by a network of perturbation-locus interactions that cannot be measured by approaches that examine perturbations or natural variation alone.

Building synthetic chromosomes from natural DNA.

De novo chromosome synthesis is costly and time-consuming, limiting its use in research and biotechnology. Building synthetic chromosomes from natural components is an unexplored alternative with many potential applications. In this paper, we report CReATiNG (Cloning, Reprogramming, and Assembling Tiled Natural Genomic DNA), a method for constructing synthetic chromosomes from natural components in yeast. CReATiNG entails cloning segments of natural chromosomes and then programmably assembling them into synthetic chromosomes that can replace the native chromosomes in cells. We use CReATiNG to synthetically recombine chromosomes between strains and species, to modify chromosome structure, and to delete many linked, non-adjacent regions totaling 39% of a chromosome. The multiplex deletion experiment reveals that CReATiNG also enables recovery from flaws in synthetic chromosome design via recombination between a synthetic chromosome and its native counterpart. CReATiNG facilitates the application of chromosome synthesis to diverse biological problems.