Optimizing Transfection of Umbilical Cord Mesenchymal Stem Cells Utilizing Minicircle plasmid/Lipofectamine LTX Complex

Lipofectamine LTX is a cationic based transfection reagent that offers high levels of different transgenes expression in a huge number of cell types in vitro utilizing an easy and rapid protocol. The level of transgenes expression depends on some experimental variables containing cell density, lipofectamine and DNA concentrations, lipofectamine -DNA complexing time, and the presence or absence of medium at the transfection time and antibiotics in medium. The significance of these factors in lipofectamine LTX based transfection will be considered. The main objective of this study is optimizing different situations for transfection ofumbilical cord mesenchymal stem cells (UC-MSCs) with lipofectamine LTX. For this purpose, UC-MSCs were transfected with lipofectamine LTX utilizing minicircle plasmid containing green fluorescent protein reporter gene. In this study, themedium existence on cell transfection efficiency as well as effects of time for culture, the ratio of lipofectamine LTX to plasmid DNA, the repetition times of transfectionand volume of medium on transfection efficacy were evaluated, and the transfection efficacies were also compared. The growth rate of MSCs wasassociated with the density of the cells. The medium changing in 4 hours after transfection certified the normal cells growth. Nevertheless, the freshness andamount of medium demonstrated no substantial effect on the cell state. The 1:2 ratio of plasmid to ipofectamine LTX (3ng:6μl), the transfection efficacy waspleasing. The transfected cells with different ratios of plasmid to lipofectamine LTX displayed substantial differences 18 h after transfection. The different conditions of MSCs transfection with lipofectamine LTX was optimized, whichpresented a reference for various adherent cells transfection.

Keywords:

Lipofectamine LTX , UC-MSCs , Transfection , Minicricle Plasmid