Synthesis and Characterization of Vitamin D₃-Functionalized Carbon Dots for Crispr/Cas9 Delivery

Abstract

Aim: To develop a novel nanovector for the delivery of genetic fragments and CRISPR/Cas9 systems in particular. Materials & methods: Vitamin D₃-functionalized carbon dots (D/CDs) fabricated using one-step microwave-aided methods were characterized by different microscopic and spectroscopic techniques. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide assay and flow cytometry were employed to determine the cell viability and transfection efficiency. Results: D/CDs transfected CRISPR plasmid in various cell lines with high efficiency while maintaining their remarkable efficacy at high serum concentration and low plasmid doses. They also showed great potential for the green fluorescent protein disruption by delivering two different types of CRISPR/Cas9 systems. Conclusion: Given their high efficiency and safety, D/CDs provide a versatile gene-delivery vector for clinical applications.

Keywords::

• carbon dots
• CRISPR/Cas9
• gene delivery
• nanovector
• vitamin D₃

Author contributions

A Hasanzadeh conceived the original idea, designed, planned and carried out the experiments, analyzed the data, designed the figures and drafted the manuscript and the supplementary information. F Radmanesh performed the cellular experiments and analyzed the data. ES Hosseini performed knock outing cellular experiments. I Hashemzadeh assisted in performing the experiments. J Kiani assisted in processing and carrying out the experiments. M Naseri assisted in performing cellular experiments. H Nourizadeh assisted in writing, language editing and proofreading the manuscript. Y Fatahi designed Scheme 1. BKY Azar extracted and purified the plasmids. BG Marani assisted in performing the experiments. A Beyzavi assisted in language editing and proofreading the manuscript. VP Mahabadi assisted in performing the experiments. M Karimi supervised the project and took the lead in writing the manuscript.

Financial & competing interests disclosure

Funding support was received from the Iran University of Medical Science (Grant 97-3-75-13318). The authors also acknowledge the Iran Science Elites Federation for funding this work. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.

Acknowledgments

The authors gratefully thank their colleagues at the Medical Nanotechnology Department and Cellular and Molecular Research Center for providing insights and expertise that greatly assisted the study.

Additional information

Funding

Funding support was received from the Iran University of Medical Science (Grant 97-3-75-13318). The authors also acknowledge the Iran Science Elites Federation for funding this work. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.