Evolutionary engineering by genome shuffling

Abstract

An upsurge in the bioeconomy drives the need for engineering microorganisms with increasingly complex phenotypes. Gains in productivity of industrial microbes depend on the development of improved strains. Classical strain improvement programmes for the generation, screening and isolation of such mutant strains have existed for several decades. An alternative to traditional strain improvement methods, genome shuffling, allows the directed evolution of whole organisms via recursive recombination at the genome level. This review deals chiefly with the technical aspects of genome shuffling. It first presents the diversity of organisms and phenotypes typically evolved using this technology and then reviews available sources of genetic diversity and recombination methodologies. Analysis of the literature reveals that genome shuffling has so far been restricted to microorganisms, both prokaryotes and eukaryotes, with an overepresentation of antibiotics- and biofuel-producing microbes. Mutagenesis is the main source of genetic diversity, with few studies adopting alternative strategies. Recombination is usually done by protoplast fusion or sexual recombination, again with few exceptions. For both diversity and recombination, prospective methods that have not yet been used are also presented. Finally, the potential of genome shuffling for gaining insight into the genetic basis of complex phenotypes is also discussed.

Tolerance engineering in bacteria for the production of advanced biofuels and chemicals

“…New technologies for genome editing (e.g., large clustered regularly interspaced palindromic repeats -interference (CRISPRi) libraries [55][56][57]) and targeted gene regulation [58] allow us to target multiple genes simultaneously and provide powerful new approaches in solvent-tolerance engineering. Genome shuffling across multiple strains with desirable phenotypes, coupled with strong selections or screens, can also be a potential approach to obtain robust strains [59].…”

Large-scale robot-assisted genome shuffling yields industrial Saccharomyces cerevisiae yeasts with increased ethanol tolerance

“…By contrast, approaches that generate artificial variation in a non-targeted fashion, for instance by means of evolutionary engineering or mutagenesis, have proven more successful to improve this complex phenotype [7]. In the last decade, genome shuffling was added to the arsenal of techniques to enhance complex phenotypes in microbes without the need for insight into the molecular mechanisms governing the trait of interest (reviewed by Biot-Pelletier and Martin [8]). Genome shuffling consists of three steps [9].…”

Microbial Propionic Acid Production

“…Compared to classical strain improvement methods such as chemical or UV mutagenesis, genome shuffling accelerates the evolutionary process by using multiple genotypes to provide an initial pool of genetic diversity, which can be refined for genomes that display desirable and diverse phenotypes. Genome shuffling combines the advantages of multi-parental crossing facilitated by DNA exchange, thus allowing the incorporation of foreign DNA [77]. Recursive genomic recombination combines classical breeding (asexual recursive mutagenesis), DNA shuffling (sexual recursive recombination) and screening for the desired phenotype and provides a feasible strategy to improve strains.…”