Emerging trends in genome integration tools for precision engineering of diverse bacterial species

TL;DR

This paper reviews tools for precise DNA insertion in bacteria, focusing on methods that work in diverse species without relying on complex host repair systems.

Contribution

The paper provides a comprehensive analysis of current and emerging genome integration systems in bacteria, emphasizing portability and precision.

Findings

Recombineering technologies have been widely used for genome integration in bacteria.

CRISPR-guided systems with integrase machinery are gaining popularity for precise DNA insertion.

Efforts to modify non-model bacterial species are increasing, expanding the scope of synthetic biology.

Abstract

The ability to precisely insert DNA payloads into a genome enables the comprehensive engineering of cellular phenotypes and the creation of new biotechnologies. To achieve such modifications, the most widely used techniques rely on a host cell’s native DNA repair mechanisms like homologous recombination, which hampers their broader use in organisms lacking these capabilities. Here, we explore the current landscape of genome integration systems with a particular focus on those that function in bacteria and are precise, self-contained, and portable, placing minimal requirements on the host cell. Through a historical analysis, we observe long-term use of recombineering technologies, a recent rise in the use of CRISPR-guided systems that consist of associated integrase machinery, and growing efforts to modify non-model organisms. Looking forward, we highlight some of the remaining challenges and how synthetic genomics may offer a way to create bacterial strains optimized for extensive long-term modification. As the field of synthetic biology sets its sights on real-world impact, the effective engineering of genomes will be critical to shaping the robust phenotypes that applications demand.

Graphical Abstract

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