Off-target genomic loci with partial complementarity to the Cx36 sgRNA

Off-target genomic loci with partial complementarity to the Cx36 sgRNA refer to unintended DNA sequences that share high sequence similarity with the guide RNA designed to target the Connexin 36 (GJD2) gene. In CRISPR-Cas9 systems, the Cas9 nuclease is guided to its target by a 20-nucleotide sgRNA; however, it can also bind and cleave at these off-target sites where the sequence is only partially complementary, leading to unintended double-strand breaks (Fu et al., 2013, Nature Biotechnology). Connexin 36 is a vital gap junction protein expressed in the central nervous system and pancreatic beta cells, where it mediates electrical synapses and synchronizes insulin release (UniProt P60591). While Cx36 is a focus for treating neurological disorders and diabetes, these off-target loci represent a significant safety risk rather than a therapeutic target. Cleavage at these sites can cause genotoxicity, including permanent mutations in unrelated genes or complex chromosomal rearrangements (Tsai et al., 2015, Nature Biotechnology). Consequently, identifying and minimizing these off-target effects is a critical requirement for the clinical translation of any Cx36-targeted gene-editing therapy. Monitoring these sites typically involves high-throughput sequencing methods to ensure the precision of the genetic intervention.

Other names

CRISPR off-target sitesGJD2 off-target lociNon-specific sgRNA binding sitesUnintended genomic cleavage sites

Unintended DNA cleavage via sequence homology-driven Cas9 recruitment