A Transfection-Free Approach of Gene Editing via a gold-based nanoformulation of the Cas9 protein

Avatar
Poster
Voices Powered byElevenlabs logo
Connected to paperThis paper is a preprint and has not been certified by peer review

A Transfection-Free Approach of Gene Editing via a gold-based nanoformulation of the Cas9 protein

Authors

Konstantinidou, S.; Lindstaedt, A.; Schmidt, T. J. N.; Nocilla, F.; Maltinti, G.; Rocco, M. A.; Landi, E.; De Carli, A.; Crucitta, S.; Lai, M.; Pistello, M.; Cappello, V.; Witt, D.; Gabellini, C.; Barski, P.; Raffa, V.

Abstract

In recent years, the CRISPR/Cas9 technology has emerged as a highly efficient tool for cell gene editing. However, the delivery of the CRISPR/Cas9 system into cells remains a significant challenge, drastically limiting in vivo gene therapy applications. In this study, we present a transfection/transduction-free tool for intracellular delivery of the Cas9:gRNA ribonucleoprotein. The Cas9 enzyme is conjugated to a 12 nm gold nanoparticle through affinity binding between the 6x His-tag of the protein and the NTA-Ni++ groups on the nanoparticles. This link chemistry allows a fine control of the density of the enzymes decorating the particle surface, the orientation of the bonding and the stability of the interaction. Importantly, the surface chemistry of this nanoformulation has been precisely engineered to modulate the cellular internalization and localization. Thanks to this approach of precision chemistry, this nanoformulation demonstrated the ability to spontaneously enter human melanoma cells as monodispersed particles that localize in cell cytoplasm, endosomes, and nucleus. It also shows effective gene editing efficiency similarly to conventional transfection tools. This gold-based formulation of Cas9 represents a ready-to-use biotech editing tool, and a promising solution for direct in vivo gene editing applications.

Follow Us on

0 comments

Add comment