Efficient Transient Expression of Plasmid DNA Using Poly (2-(N, N-Dimethylamino) Ethyl Methacrylate) in Plant Cells

Zishuai An; Bing Cao; Junzhe Zhang; Baihong Zhang; Chengqian Zhou; Xianglong Hu; Wenli Chen

doi:10.3389/fbioe.2022.805996

Efficient Transient Expression of Plasmid DNA Using Poly (2-(N, N-Dimethylamino) Ethyl Methacrylate) in Plant Cells

Front Bioeng Biotechnol. 2022 Feb 22:10:805996. doi: 10.3389/fbioe.2022.805996. eCollection 2022.

Authors

Zishuai An^{1

2

3}, Bing Cao^{1

2

3}, Junzhe Zhang^{1

2

3}, Baihong Zhang^{1

2

3}, Chengqian Zhou⁴, Xianglong Hu^{1

2

3}, Wenli Chen^{1

2

3}

Affiliations

¹ MOE Key Laboratory of Laser Life Science and Institute of Laser Life Science, College of Biophotonics, South China Normal University, Guangzhou, China.
² Guangdong Provincial Key Laboratory of Laser Life Science, College of Biophotonics, South China Normal University, Guangzhou, China.
³ Guangzhou Key Laboratory of Spectral Analysis and Functional Probes, College of Biophotonics, South China Normal University, Guangzhou, China.
⁴ Neuroscience Laboratory, Hugo Moser Research Institute at Kennedy Krieger, Baltimore, MD, United States.

Abstract

Nanomaterials have been widely studied for their potential to become the new generation of nanocarriers in gene transfection, yet it remains still difficult to apply them efficiently and succinctly to plant cells. Poly (2-(N,N-dimethylamino) ethyl methacrylate) (PDMAEMA), which possesses temperature and pH dual-sensitivity, has largely been applied in animal cells, but it is rarely involved in plant cells. As a proof of concept, PDMAEMA as a gene carrier is incubated with plasmid GFP (pGFP) to explore its transfection ability in plants, and cationic polymer polyethylenimine (PEI) is used as a control. pGFP was efficiently condensed into the nanostructure by electrostatic interactions at an N/P (amino group from cationic polymers/phosphate group from plasmid DNA (pDNA)) ratio of 15; after complexation into nanocarriers, pGFP was protected from endonuclease degradation according to the DNase I digestion assay. After incubation with protoplasts and leaves, GFP was observed with confocal microscopy in plant cells. Western blot experiments confirmed GFP expression at the protein level. Toxicity assay showed PDMAEMA had a lower toxicity than PEI. These results showed that transient expression of pGFP was readily achieved in Arabidopsis thaliana and Nicotiana benthamiana. Notably, PDMAEMA showed lower cytotoxicity than PEI upon incubation with Nicotiana benthamiana leaves. PDMAEMA exhibited great potency for DNA delivery in plant cells. This work provides us with new ideas of more concise and more effective methods for plant transformation.

Keywords: N-dimethylamino) ethyl methacrylate) (PDMAEMA); gene delivery; gene transfection; plant cells; poly (2-(N; polyethylenimine (PEI).