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Cell-Penetrating Peptide Delivery for CRISPR Applications
Abhik Paul1, Hye Young Lee1, Jan Tuma2
1The Department of Cellular and Integrative Physiology, The University of Texas Health Science Center at San Antonio, San Antonio, TX, USA.
Abstract:
Cell-penetrating peptides (CPPs) provide a modular nonviral strategy for intracellular delivery of CRISPR/Cas9 components, particularly preassembled Cas9 ribonucleoprotein (RNP) complexes. This chapter summarizes key principles of CPP-mediated Cas9 RNP delivery, including noncovalent complexation and covalent CPP-Cas9 conjugation strategies. A representative protocol is provided for the preparation and evaluation of Cas9-K5 RNP, including in vitro uptake and reporter-editing assays, nucleofection-based functional controls, local intracranial delivery, immunofluorescence staining, and quantitative analysis of reporter activation in brain tissue.