Abstract:

Objective　To determine the optimized condition under which BMP expression vector will be constructed to transfect bone marrowstromal cells (MSCs), plasmid vector coding enhanced green fluorescence protein (EGFP) gene pEGFPwas transferred into MSCs. The transfer efficiency and transient expression were subsequently tested.Methods　pEGFP plasmid was amplified and tested by an enzyme cutting technique in vitro. MSCs, which were initially obtained from the bone marrow of rab- bits, were culturedin vitroand transferred with pEGFP by means of lipofectamine media methods. The ratio of plasmid and lipo- fectamine was varied according to the experiment design. Transfer efficiency and transient expressionwere evaluated by fluorescent microscopy.Results　Transfer efficiencywas correlatedwith the ratio of plamid and lipofectamine. The expression of EGFP began in 24 hours after transferring, reached maximum in 48-72 hours and decreased in 1 week, however there remained a weak expres- sion for more than 3 weeks.Conclusion　The efficiency of transferring pEGFP intoMSCs could achieve to 30% with proper ratio of plamid and lipofactamine. pEGFP was an ideal transient expression vector forMSCs gene transference.

Key words: enhanced green fluorescent protein, marrow stromal cells, gene transference, lipofectamine