CRISPR/Cas9 Genome Editing Using Gold-Nanoparticle-Mediated Laserporation

Publikation: Beitrag in FachzeitschriftArtikelForschungPeer-Review

Autoren

  • Berislav Bošnjak
  • Marc Permanyer
  • Maya K. Sethi
  • Melanie Galla
  • Tobias Maetzig
  • Dag Heinemann
  • Stefani Willenzon
  • Reinhold Förster
  • Alexander Heisterkamp
  • Stefan Kalies

Organisationseinheiten

Externe Organisationen

  • Medizinische Hochschule Hannover (MHH)
  • Laser Zentrum Hannover e.V. (LZH)
  • NIFE- Niedersächsisches Zentrum für Biomedizintechnik, Implantatforschung und Entwicklung
  • REBIRTH Forschungszentrum für translationale regenerative Medizin
Forschungs-netzwerk anzeigen

Details

OriginalspracheEnglisch
Aufsatznummer1700184
FachzeitschriftAdvanced Biosystems
Jahrgang2
Ausgabenummer11
Frühes Online-Datum29 Mai 2018
PublikationsstatusVeröffentlicht - 9 Nov. 2018

Abstract

Engineered nucleases hold large potential for future gene therapy applications. An obstacle hampering their applications are delivery methods bearing efficiency, throughput, and viability of target cells. How this limitation can be overcome via gold-nanoparticle-mediated (GNOME) laserporation is demonstrated. It employs a picosecond laser setup and 200 nm gold nanoparticles, and its full capacity with CRISPR/Cas9 delivery is demonstrated. 70 kDa dextrans are utilized to probe delivery in adherent SC1 cells. Afterward, GNOME laserporation is used for transfection of crRNA:tracrRNA targeting the mouse CCR7 (mCCR7) into SpCas9 (Streptococcus pyogenes Cas9) and mCCR7 co-expressing SC1 cells. Finally, ribonucleoprotein particles consisting of mCCR7 crRNA:tracrRNA and SpCas9 endonuclease are transfected into SC1 cells not expressing SpCas9. Gene knockout efficiencies of up to 65% are detected in the GNOME laserporated cells. To validate the simplicity of the approach, the same treatment parameters are used to successfully knock out CXCR3 in 25% of GNOME laserporated activated mouse CD8+ T cells. In conclusion, this is the first demonstration of the unique combination of nanotechnology and laser irradiation for gene editing via engineered nucleases.

ASJC Scopus Sachgebiete

Zitieren

CRISPR/Cas9 Genome Editing Using Gold-Nanoparticle-Mediated Laserporation. / Bošnjak, Berislav; Permanyer, Marc; Sethi, Maya K. et al.
in: Advanced Biosystems, Jahrgang 2, Nr. 11, 1700184, 09.11.2018.

Publikation: Beitrag in FachzeitschriftArtikelForschungPeer-Review

Bošnjak, B, Permanyer, M, Sethi, MK, Galla, M, Maetzig, T, Heinemann, D, Willenzon, S, Förster, R, Heisterkamp, A & Kalies, S 2018, 'CRISPR/Cas9 Genome Editing Using Gold-Nanoparticle-Mediated Laserporation', Advanced Biosystems, Jg. 2, Nr. 11, 1700184. https://doi.org/10.1002/adbi.201700184
Bošnjak, B., Permanyer, M., Sethi, M. K., Galla, M., Maetzig, T., Heinemann, D., Willenzon, S., Förster, R., Heisterkamp, A., & Kalies, S. (2018). CRISPR/Cas9 Genome Editing Using Gold-Nanoparticle-Mediated Laserporation. Advanced Biosystems, 2(11), Artikel 1700184. https://doi.org/10.1002/adbi.201700184
Bošnjak B, Permanyer M, Sethi MK, Galla M, Maetzig T, Heinemann D et al. CRISPR/Cas9 Genome Editing Using Gold-Nanoparticle-Mediated Laserporation. Advanced Biosystems. 2018 Nov 9;2(11):1700184. Epub 2018 Mai 29. doi: 10.1002/adbi.201700184
Bošnjak, Berislav ; Permanyer, Marc ; Sethi, Maya K. et al. / CRISPR/Cas9 Genome Editing Using Gold-Nanoparticle-Mediated Laserporation. in: Advanced Biosystems. 2018 ; Jahrgang 2, Nr. 11.
Download
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AU - Bošnjak, Berislav

AU - Permanyer, Marc

AU - Sethi, Maya K.

AU - Galla, Melanie

AU - Maetzig, Tobias

AU - Heinemann, Dag

AU - Willenzon, Stefani

AU - Förster, Reinhold

AU - Heisterkamp, Alexander

AU - Kalies, Stefan

N1 - © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

PY - 2018/11/9

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N2 - Engineered nucleases hold large potential for future gene therapy applications. An obstacle hampering their applications are delivery methods bearing efficiency, throughput, and viability of target cells. How this limitation can be overcome via gold-nanoparticle-mediated (GNOME) laserporation is demonstrated. It employs a picosecond laser setup and 200 nm gold nanoparticles, and its full capacity with CRISPR/Cas9 delivery is demonstrated. 70 kDa dextrans are utilized to probe delivery in adherent SC1 cells. Afterward, GNOME laserporation is used for transfection of crRNA:tracrRNA targeting the mouse CCR7 (mCCR7) into SpCas9 (Streptococcus pyogenes Cas9) and mCCR7 co-expressing SC1 cells. Finally, ribonucleoprotein particles consisting of mCCR7 crRNA:tracrRNA and SpCas9 endonuclease are transfected into SC1 cells not expressing SpCas9. Gene knockout efficiencies of up to 65% are detected in the GNOME laserporated cells. To validate the simplicity of the approach, the same treatment parameters are used to successfully knock out CXCR3 in 25% of GNOME laserporated activated mouse CD8+ T cells. In conclusion, this is the first demonstration of the unique combination of nanotechnology and laser irradiation for gene editing via engineered nucleases.

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