A patent-based consideration of latest platforms in the art of directed evolution: a decade long untold story

Directed (or in vitro) evolution of proteins and metabolic pathways requires tools for creating genetic diversity and identifying protein variants with new or improved functional properties. Besides simplicity, reliability, speed, versatility, universal applicability and economy of the technique, the new science of synthetic biology requires improved means for construction of smart and high-quality mutant libraries to better navigate the sequence diversity. In vitro CRISPR/Cas9-mediated mutagenic (ICM) system and machine-learning (ML)-assisted approaches to directed evolution are now in the field to achieve the goal. This review describes the gene diversification strategies, screening and selection methods, in silico (computer-aided), Cas9-mediated and ML-based approaches to mutagenesis, developed especially in the last decade, and their patent position. The objective behind is to emphasize researchers the need for noting which mutagenesis, screening or selection method is patented and then selecting a suitable restriction-free approach to sequence diversity. Techniques and evolved products subject to patent rights need commercial license if their use is for purposes other than private or experimental research.