CRISPR with a Happy Ending: Non‐Templated DNA Repair for Prokaryotic Genome Engineering

The exploration of microbial metabolism holds enormous power to help society shift towards an environmentally sustainable economy and to tackle a plethora of problems related to the burdens of human consumption. Microbial cell factories have the potential to catalyze a wide range of processes which are currently either unavailable, unsustainable and/or inefficient. The metabolism of microorganisms can be optimized and further expanded using genetic engineering tools, like the CRISPR-Cas systems. These tools have revolutionized the field of biotechnology, as they greatly facilitate the genetic optimization of organisms from all domains of life. These and other nucleases mediate double-strand DNA breaks, which must be repaired to prevent cell death. These breaks can be repaired in prokaryotes through either homologous recombination, when a DNA repair template is available, or through template-independent end joining, of which two major pathways are known. These end joining pathways depend on different sets of proteins and mediate DNA repair with different outcomes. Understanding these DNA repair pathways can therefore be advantageous to steer the results of genome engineering experiments. In this review, we discuss different strategies for the genetic engineering of prokaryotes through the exploitation of either the non-homologous end joining (NHEJ) or the alternative end joining (AEJ) DNA repair pathways, which are independent of exogenous DNA repair templates. This article is protected by copyright. All rights reserved.