Cas12b target DNA sequence (Protospacer)

The target DNA sequence complementary to the Cas12b guide RNA, located adjacent to a T-rich protospacer adjacent motif (PAM), serves as the specific genomic or extrachromosomal site for CRISPR-Cas12b-mediated intervention (Shmakov et al., 2015). Cas12b, a Type V-B CRISPR-associated endonuclease, utilizes a guide RNA to recognize and bind this sequence through Watson-Crick base pairing after initial PAM recognition (Liu et al., 2017). Upon binding, Cas12b induces a double-strand break in the target DNA, which can be leveraged for gene knockout, insertion, or correction in therapeutic contexts (Strecker et al., 2019). Beyond direct genome editing, the recognition of this target sequence triggers the collateral non-specific single-stranded DNA (ssDNA) cleavage activity of Cas12b, a property widely exploited in highly sensitive molecular diagnostic assays for detecting pathogens or genetic mutations (Li et al., 2018). Therapeutic strategies targeting these sequences aim to treat monogenic disorders, such as sickle cell disease or cystic fibrosis, by precisely modifying the underlying genetic cause. However, the primary challenge remains ensuring high specificity to avoid off-target cleavage at similar DNA sequences, which could lead to unintended genomic instability.

Other names

ProtospacerCas12b target siteCRISPR-Cas12b recognition sequenceC2c1 target DNA

The Cas12b protein, guided by a specific RNA sequence, recognizes a T-rich PAM and binds to the complementary target DNA sequence, leading to site-specific double-stranded DNA cleavage and activation of collateral trans-cleavage activity.