Optimizing A Lipocomplex-Based Gene Transfer Method into HeLa Cell Line

Abstract

One of the most significant steps in gene expression studies is transferring genes into cell cultures. Despite there are different methods for gene delivery such as viral and non-viral producers, some cationic lipid reagents have recently developed to transfect into mam- malian cell lines. The main aim of this study was optimizing and improving lipocomplex based transient transfection procedures into HeLa cell line which is being used widely as a typical cell in biological studies. This study was an experimental research. In this work, pCMV β-Gal DNA plasmid was used as a reporter DNA for determining the rate of gene transfection into HeLa cells. To accomplish the highest gene delivery into HeLa cells, optimizing experiments were car- ried out in different volumes of FuGENE-HD, Lipofectamine(TM)2000 and X-tremeGENE. Also, we investigated tranasfection efficiency in presence of various cell densities of HeLa cells. Then, transfection efficiency and cell toxicity were measured by beta gal staining and trypan blue methods, respectively. Using FuGENE-HD in volume of 4µl along with 10(5) HeLa cells, transfection efficiency was higher (43.66 ± 1.52%) in comparison with the cationic lipids Lipofectamine(TM)2000 and X-tremeGENE. In addition, the rate of cell toxicity in presence of FuGENE-HD was less than 5%. In summary, the cationic lipid FuGENE-HD indicates a suitable potential to transfer DNA into HeLa cells and it can be an efficient reagent for gene delivery for HeLa cells in vitro. Moreover, it is worth designing and optimizing gene transfer experiments for other cell lines with FuGENE-HD due to its low toxicity and high efficiency.

Keywords: HeLa Cells; Lipids; Transfection.

Fig 1

The best result for transfection was achieved in presence of 4 μl FuGENE-HD. Also, increasing the volume of FuGENE-HD, not only did not lead to increase of transfection efficiency, but it also caused more cell toxicity.

Fig 2

The graph shows optimizing transfection efficiency in different cell densities while the volume of Lipofectamine^TM2000 fixed at 4 μl.

Fig 3

In this comparative graph, the percentage of transfection in presence different cationic lipid reagents has been shown. The rate of transfection of HeLa cells in presence of FuGENE-HD, Lipofectamine^TM2000 and X-trem- GENE was 43.66 ± 1.52 %, 31.66 ± 2.5% and 4.33 ± 1.15%, respectively.

Fig 4

These pictures show the condition of cell viability and the level of transfection efficiency into transfected HeLa cells under microscope in presence of A. FuGENE-HD, B. Lipofectamine^TM2000 and C. X-tremGENE. The blue cells express beta galactosidase activity.