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Efficient Intracellular Delivery of Native Proteins

Author:
D’Astolfo, Diego S., Pagliero, Romina J., Pras, Anita, Karthaus, Wouter R., Clevers, Hans, Prasad, Vikram, Lebbink, Robert Jan, Rehmann, Holger, Geijsen, Niels
Source:
Cell 2015 v.161 pp. 674-690
ISSN:
0092-8674
Subject:
DNA, gene targeting, messenger RNA, proteins, sodium chloride
Abstract:
Modulation of protein function is used to intervene in cellular processes but is often done indirectly by means of introducing DNA or mRNA encoding the effector protein. Thus far, direct intracellular delivery of proteins has remained challenging. We developed a method termed iTOP, for induced transduction by osmocytosis and propanebetaine, in which a combination of NaCl hypertonicity-induced macropinocytosis and a transduction compound (propanebetaine) induces the highly efficient transduction of proteins into a wide variety of primary cells. We demonstrate that iTOP is a useful tool in systems in which transient cell manipulation drives permanent cellular changes. As an example, we demonstrate that iTOP can mediate the delivery of recombinant Cas9 protein and short guide RNA, driving efficient gene targeting in a non-integrative manner.
Agid:
5517110