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New CRISPR Editor The evoCasΦ2, a revised version of the compact CRISPR editor CasΦ2, edited DNA in human cells up to 70 times more actively than the original enzyme. On August 13, a preprint was published on bioRxiv about evoCasΦ2, a version of CasΦ2 with six amino acid substitutions. The authors compared it to the original enzyme and two versions created by rational design on four genomic sites in the human cell line HEK293T. The CasΦ2 is a CRISPR protein that finds a target DNA site by guide RNA and cuts it. It has 757 amino acids, requiring less space in the delivery system than the larger Cas9. The original CasΦ2 barely edited DNA in mammalian cells, and successful variants were difficult to distinguish from random noise. To address this, the authors built EPICA.2, a two-step directed evolution system. They created many slightly modified CasΦ2 variants and first selected them in yeast: DNA cutting restored gene function, allowing the cell to grow. Four rounds of mutation and re-selection gathered a library of more active variants. The library was then transferred to human cells, where a gene for green fluorescent protein was shifted and non-functional. CasΦ2 cutting sometimes restored the gene's correct reading, the protein began to glow, and the authors selected such cells. Long reading of each DNA molecule with an individual tag preserved the set of substitutions within one variant, leading to the discovery of evoCasΦ2 with six substitutions. The effect is created by the combination of these six substitutions. On a control site, no single substitution repeated it, and removal of any from the full version reduced activity. On four native sites in the HEK293T cell genome, evoCasΦ2 edited DNA up to 70 times more actively than the original CasΦ2 and outperformed two versions created by rational design. On individual targets, the fraction of edited DNA remained below 5%. EPICA.2 continues the first EPICA, which in 2024 gave a variant of another compact nuclease, CjCas9, with activity up to 12 times higher than the original enzyme on endogenous sites in human cells, as reported in Nature Aging, July 2026. 🔗 Read original →