CS-Cells: A CRISPR-Cas12 DNA Device to Generate Chromosome-Shredded Cells for Efficient and Safe Molecular Biomanufacturing

Abstract

Synthetic biology holds great promise for translating ideas into products to address the grand challenges facing humanity. Molecular biomanufacturing is an emerging technology that facilitates the production of key products of value, including therapeutics and select chemical compounds. Current biomanufacturing technologies require improvements to overcome limiting factors, including efficient production, cost, and safe release; therefore, developing optimum chassis for biomolecular manufacturing is of great interest for enabling diverse synthetic biology applications. Here, we harnessed the power of the CRISPR-Cas12 system to design, build, and test a DNA device for genome shredding, which fragments the native genome to enable the conversion of bacterial cells into nonreplicative, biosynthetically active, and programmable molecular biomanufacturing chassis. As a proof of concept, we demonstrated the efficient production of green fluorescent protein and violacein, an antimicrobial and antitumorigenic compound. Our CRISPR-Cas12-based chromosome-shredder DNA device has built-in biocontainment features providing a roadmap for the conversion of any bacterial cell into a chromosome-shredded chassis amenable to high-efficiency molecular biomanufacturing, thereby enabling exciting and diverse biotechnological applications.