Figure 3.

CRISPR - A versatile toolkit for the targeted deletion, rewriting and insertion of novel genetic information. Continuous iterations of technological innovation have considerably extended the scope and versatility of CRISPR-based gene editing strategies. Today, CRISPR genome editing encompasses a wide spectrum of diverse applications including the direct rewriting of genetic code, modulation of gene expression, and post-transcriptional RNA editing. While the most widely known CRISPR-Cas9 system and others rely on inducing DNA double-strand breaks (DSBs) and carry the potential risk of introducing deleterious effects arising from off-target cleavages (A), other CRISPR-based editing tools, including base editing (105, 106) and prime editing (107), allow to rewrite the genetic code without the requirement for DNA double-strand cleavages and, hence, present a potentially safer alternative (B). Over time, the possibilities to insert novel genetic material have expanded dramatically with the most recent advances including Twin prime editing (108) and CRISPR-associated transposase (CAST)-editing (109) now allowing large transgenic inserts in the range of up to 10-40 kb length. Besides editing the genetic code, powerful tools now exist which allow to tune transcriptional activation in a targeted manner. CRISPRa and CRISPRi rely on the site-directed delivery of transcriptional effector molecules to activate or repress gene expression (110–113). More profound epigenetic modulation can be achieved by tethering catalytically dead Cas9 to methyltransferases and acetyltransferases to alter the global methylome and edit histone modifications (114–124). Created with BioRender.com.