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Proc Natl Acad Sci U S A. 2019 Apr 16;116(16):7899-7904. doi: 10.1073/pnas.1818553116. Epub 2019 Mar 28.

Nontoxic nanopore electroporation for effective intracellular delivery of biological macromolecules.

Author information

1
Department of Chemistry, University of California, Berkeley, CA 94720.
2
Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720.
3
Department of Materials Science and Engineering, University of California, Berkeley, CA 94720.
4
Department of Molecular and Cellular Physiology, Stanford University, Stanford, CA 94705.
5
Howard Hughes Medical Institute, University of California, Berkeley, CA 94720.
6
Innovative Genomics Institute, University of California, Berkeley, CA 94720.
7
Molecular Biophysics & Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720.
8
California Institute for Quantitative Biosciences, Berkeley, CA 94720.
9
Department of Chemistry, University of California, Berkeley, CA 94720; p_yang@berkeley.edu.

Abstract

We present a simple nanopore-electroporation (NanoEP) platform for delivery of nucleic acids, functional protein, and Cas9 single-guide RNA ribonucleoproteins into both adherent and suspension cells with up to 80% delivery efficiency and >95% cell viability. Low-voltage electric pulses permeabilize a small area of cell membrane as a cell comes into close contact with the nanopores. The biomolecule cargo is then electrophoretically drawn into the cells through the nanopores. In addition to high-performance delivery with low cell toxicity, the NanoEP system does not require specialized buffers, expensive materials, complicated fabrication processes, or cell manipulation; it simply consists of a generic nanopore-embedded water-filter membrane and a low-voltage square-wave generator. Ultimately, the NanoEP platform offers an effective and flexible method for universal intracellular delivery.

KEYWORDS:

electroporation; genome engineering; intracellular delivery; nanopore; nanotechnology

PMID:
30923112
DOI:
10.1073/pnas.1818553116

Conflict of interest statement

Conflict of interest statement: Y.C., E.M., J.A.D., and P.Y. are inventors on patent applications (filed by the University of California, Berkeley) related to the nanopore electroporation systems and uses thereof. J.A.D. is a cofounder of Caribou Biosciences, Editas Medicine, Intellia Therapeutics, Scribe Therapeutics, and Mammoth Biosciences and is a member of the board of directors of Driver and Johnson & Johnson and a member of the scientific advisory boards for Caribou Biosciences, Intellia Therapeutics, eFFECTOR Therapeutics, Scribe Therapeutics, Synthego, Metagenomi and Inari. Y.C., S.C.-B., P.Y., and Bozhi Tian are coauthors on a 2018 review article.

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