Assessment of two transfection techniques in the switch of mir-۱۹۲ expressing plasmid in HDFand NALM-۶ cell strains

Introduction: Due to the recent developments in the field of biotechnology and genetics, transfection hasbecome popular to study cellular processes and molecular mechanisms of diseases. During Transfection,different types of nucleic acids such as DNA, RNA, as well as small and non-coding RNAs such as siRNA,shRNA, miRNA can be transferred to mammalian cells. Several studies have shown that mir-۱۹۲ plays a rolein cancer progression. We have various transfection methods including Polycation Polyethylene (PEI), andLipofectamine ۳۰۰۰. PEI is a synthetic polymer with an extremely high positive charge density, and stronglybinds to the negatively charged DNA, creating a net cationic charge and allowing the DNA to enter cells.Lipofectamine form a monolayer liposomal structure with a positive surface charge and combine with nucleicacids to form a transfection complex. Human dermal fibroblasts (HDF) are stromal adherent cells that providethe majority of the structural framework of almost all tissue types. NALM-۶ is a B-cell progenitor leukemia cellline that is suspended. Our aim of this study is to compare different transfection methods for the transferPLentiIII-miR-۱۹۲-GFP plasmid in HDF and NALM-۶ cell lines.Methods: In this study, bacterial culture were performed in order to prepare PLentiIII-miR-۱۹۲-GFP, and thenthe plasmid was extracted. HDF and NALM-۶ cells were cultured for growth and maintenance in mediumcontaining FBS, penicillin and streptomycin respectly. PEI and lipofectamine ۳۰۰۰ methods were used totransfer ۲μg PLentiIII-miR-۱۹۲-GFP into the cells. Then, intransfected cells, the expression of GFP tag in eachof the methods was measured by flow cytometry device. The steps of cDNA synthesis were done from mRNAand finally the expression of mir-۱۹۲ was measured by qRT-PCR.Results: For HDF cells, in the PEI method, the transfection rate was ۲۰-۳۰%, and in lipofection, the rate was۵-۷%. For NALM-۶, in the lipofection۳۰۰۰ method, the transfection rate was ۱۵%, and in the other method itwas less than ۱۰%.Conclusion: Regarding the advantages and disadvantages of the methods, the PEI method is easy and providestransfection efficiency and high titer. The main disadvantage of PEI is its high cationic charge density andnonbiodegradability. In the lipofection method, it works on many types of cells, including cultured nerve cells.It allows transfection reactions to be performed in ۳۰ minutes. Its disadvantages include low transfection,inability to target specific cells, short half-life, and toxicity caused by positively charged lipids. However, adifferent method should be set up for each cell depending on its nature.