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Transfection of nucleic acids is a popular technology in oncology research and gene therapy. Transfection efficiency is the major limitation impacting efficacy. However, there is little research on the molecular determinants of transfection efficiency.We investigated the biological effect of lipofection of non-operative siRNA, and identified a gene cluster relevant to efficiency. FITC-labeled siRNA was transfected into the HEK293 cells using nine distinct transfection reagents under respective optimized conditions. Transfection efficiencies of the reagents were determined by cytometric detection of intracellular FITC. Cellular damage by each reagent was determined by MTT or trypan blue staining. To identify the cellular determinants for transfection efficiency and cell damage, we performed gene expression analysis using GeneChip (U133 ver.2). The gene expression profile was analyzed by BRB ArrayTools. The transfection efficacy of the eight reagents was generally ∼60-80%, but one lipofectant showed low efficiency (∼20%). No correlation was observed between the transfection efficiencies of these reagents and cellular damage. Some membrane charge-related genes, such as ATP10D and SFXN1, were selected as genes specifically regulated by lipofection (by t-test) and their expression changes were confirmed by RT-PCR. These genes were suggested to correlate with trasfection efficiencies. Biological confirmations of some of the candidate genes are currently underway. The function of each candidate is being examined by employing transfection techniques. Using these biomarkers to assess transfection efficiency, it may be possible to predict the transfection efficiency of tumors.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]