Effect of Cell Passage Time on the Electrotransfection Efficiency
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ELL BIOLOGY
Effect of Cell Passage Time on the Electrotransfection Efficiency Sonam Chopraa, *, Paulius Ruzgysa, **, Martynas Maciulevičiusa, ***, and Saulius Šatkauskasa, **** a
Biophysical Research Group, Faculty of Natural Sciences, Vytautas Magnus University, Vileikos str. 8, LT 44404, Kaunas, Lithuania *e-mail: [email protected] **e-mail: [email protected] ***e-mail: [email protected] ****e-mail: [email protected] Received December 16, 2019; revised March 2, 2020; accepted March 2, 2020
Abstract—Gene electrotransfer is an effective and promising gene delivery technique in clinical applications, such as DNA vaccination and gene therapy. An improved gene therapy protocol depends on the the proper establishment of the gene transfer method. Electroporation has been widely employed in in vitro and in vivo protocols, and increaing its transfection efficiency has been the field of research. In order to achieve the the maximal introduction of plasmid DNA into cells with optimal cell viability, electro transfection conditions for every single cell type should be determined individually. In this work, the effect of cell passage time on the electrotransfection efficiency of CHO cells is determined. The selected cell passage times of 24 and 48 h prior to the electroporation are considered for the analysis. It is shown that electrotransfection efficiency with all plasmid concentrations significantly differs when comparing 24 and 48 h cell passage times. However, only slight change in the cell viability is observed at 24 and 48 h of cell passage times. Keywords: plasmid DNA, electroporation, electrotransfection, optimization, cell passage time DOI: 10.1134/S1062359020550014
INTRODUCTION Gene therapy is the emerging technology to treat or prevent disease by replacing a defective or missing gene (Wirth et al., 2013). A wide range of candidate genes for gene therapy have been divulged but very few have turned into target therapies because of the poor delivery of the nucleic acid (Rao and Zacks, 2014; Mostaghaci et al., 2016). Many viral and non-viral gene delivery methods have been used in cell transfection. Viral vectors have been thoroughly investigated and demonstrated to have high transfection efficiencies. However, they have potential risks such as immunogenicity, insertional mutagenesis, oncogenicity, etc. (Mulligan, 1993; Li and Huang, 2000; Kohn et al., 2003). Non-viral (chemical or physical) gene delivery approaches have been developed to overcome the inherent problems of viral gene vectors. Among physical approaches, electroporation or electropermeabilization is one of the most promising methods for gene electrotransfer (Chopra and Satkauskas, 2018; Neumann et al., 1982; Rols et al., 1998; Satkauskas et al., 2002). Electroporation technique is based on the application of external electric field that permeabilizes the cell membrane by inducing transmembrane potential (Bonnafous et al., 1999; Neumann et al., 1999; Gehl, 2003; Gift and Weaver, 2000; Somiari et al., 2000; Phez et al., 2005). W
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