Fig. 5: Strategies for enhanced genomic resolution.

CRISPR–dCas9-based imaging techniques have been utilized to improve genomic resolution, enabling the visualization of nonrepetitive genomic regions and SNP sites. a Efficient visualization of nonrepetitive sequences. Early approaches demonstrated the feasibility of visualizing nonrepetitive sequences by introducing multiple sgRNAs (~30 sgRNAs) to a confined genomic region. Signal amplification strategies were subsequently adopted to reduce the number of sgRNAs required. The use of multiple MS2 stem loops in sgRNAs or MB-based FRET approaches enabled visualization of the same genomic region with only four or three sgRNAs, respectively. Introducing PUF or foldon allowed visualization of target sites with just a single sgRNA. b SNP site detection in living cells. The PAM sequence plays a critical role in target specificity, with point mutations in the second or third PAM positions effectively preventing RNP complex binding to the target gene. By designing sgRNAs to position SNP sites within PAM regions, allele-specific SNP mutations can be visualized in living cells.