Abstract
The development of CRISPR-Cas9 mediated gene editing technology is revolutionizing molecular biology, biotechnology, and medicine. However, as with other nucleic acid technologies, CRISPR would greatly benefit from chemical modifications that optimize delivery, activity, and specificity of gene editing. Amide modifications at certain positions of short interfering RNAs have been previously shown to improve their RNAi activity and specificity, which motivated the current study on replacement of selected internucleoside phosphates of CRISPR RNAs with amide linkages. Herein, we show that amide modifications did not interfere with CRISPR-Cas9 activity when placed in the protospacer adjacent motif (PAM) distal region of CRISPR RNAs. In contrast, modification of the seed region led to a loss of DNA cleavage activity at most but not all positions. These results are encouraging for future studies on amides as backbone modifications in CRISPR RNAs.
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