Expanding the Genome Editing Toolbox: Cas12a Unlocks New Areas of Yeast DNA

For years, Saccharomyces cerevisiae—common baker’s yeast—has been a vital organism for biotechnology and basic research. While the revolutionary CRISPR/Cas9 system has allowed scientists to edit yeast genomes rapidly, it faces a significant limitation: Cas9 is selective. It specifically targets DNA sequences rich in guanine, relying on specific markers known as protospacer-adjacent motifs (PAMs) to bind and cut. This leaves large sections of the genome, particularly those rich in thymine, difficult to reach.

To bridge this gap, researchers have established a complementary system using CRISPR/Cas12a. Unlike its predecessor, Cas12a is designed to target T-rich PAM sequences, effectively unlocking previously inaccessible genomic regions. Through rigorous benchmarking, the team identified a specific variant, improved LbCas12a (impLbCas12a), as the most efficient tool for the job. It demonstrates high editing purity and precise control over the editing window.

Crucially, this new system supports multiplexing, meaning it can install multiple genetic variants simultaneously via a homology-integrated array. This method allows for targeted 'saturation mutagenesis', a technique where scientists create comprehensive libraries of mutations to discover novel functions. By combining this new T-rich targeting tool with the established G-rich targeting of Cas9, geneticists can now interrogate a much broader genomic sequence space than ever before.