A summary of emerging technologies in the genome editing field. DNA polymerase editors: this technology combines Cas9 nickases with DNA polymerases and tethering of a single-stranded DNA template, for example, using an HUH endonuclease. A key difference from prime editing lies in its use of DNA polymerase rather than reverse transcriptase and the delivery of the DNA template intrans. CRISPR-associated transposons: these naturally occurring mobile genetic elements utilize CRISPR effector complexes in conjunction with transposase proteins for RNA-guided transposition to insert long DNA sequences into specific genomic sites. Engineered CRISPR integrases: these technologies are based on combining prime editors with site-specific serine recombinases. The prime editing initially introduces a recombinaseattsite at the target DNA location, subsequently enabling recombinase-catalyzed insertion of large DNA payloads. Target-primed reverse transcription: this process involves fusing nickase Cas9 with non-long terminal repeat (non-LTR) retrotransposon-derived reverse transcriptases and RNAs. It operates by nicking the target DNA to generate a free 3 end to prime reverse transcription of the retrotransposon-associated RNA, resulting in targeted DNA insertion. Epigenetic editors: fusions of deactivated dCas9 with DNA methylases and histone-modification enzymes enable targeted chromatin modifications at specific genomic locations, leading to the heritable repression of gene expression (CRISPRoff) without altering the underlying DNA sequence. Gene reactivation (CRISPRon) involves targeting repressed genes using Cas9 fusions with DNA demethylases and transcriptional activator domains. Artificial intelligence in gene editing: AI is making significant inroads inde novoprotein and guide design, as well as in computational prediction of off-target sites and editing outcomes.
