Off-target genomic adenine

Off-target genomic adenine within double-stranded DNA at near-match protospacer sites with NRCH PAMs refers to unintended sites of genetic modification by Adenine Base Editors (ABEs) utilizing engineered Cas9 variants. These variants, such as SpCas9-NRCH, are designed to recognize a broader range of Protospacer Adjacent Motifs (PAMs) beyond the standard NGG, specifically NRCH (where R is A/G and H is A/C/T) (Miller et al., 2020, Nature Biotechnology). When an ABE is directed to a target site, it may also bind to and deaminate adenines at "near-match" genomic locations that share high sequence homology with the guide RNA (Gaudelli et al., 2017, Nature). This process converts adenine to inosine, which is subsequently treated as guanine by DNA polymerase, resulting in permanent A•T-to-G•C transitions (Rees & Liu, 2018, Nature Reviews Genetics). Such off-target activity is a primary safety concern in genome editing, as it can lead to deleterious mutations, loss of tumor suppressor function, or activation of oncogenes (Zuo et al., 2019, Science). Consequently, identifying and quantifying these sites using high-throughput sequencing methods like GUIDE-seq or DISCOVER-seq is essential for the clinical translation of base editing technologies (Wienert et al., 2019, Science).

Other names

Off-target genomic adenine within double-stranded DNA at near-match protospacer sites with NRCH PAMsOff-target A•T-to-G•C conversionNon-specific adenine deaminationSpCas9-NRCH off-target siteNear-match protospacer adenine

Unintended adenosine deamination by a CRISPR-guided base editor at non-target genomic loci, leading to A-to-G transitions.