Specific inhibition of green fluorescence protein by small hairpin RNA (shRNA) in human glioma BT325 cell lines

Objective To investigate whether the green fluorescence protein (GFP) specific short hairpin RNA (shRNA) can inhibit the expression of GEP gene in human glioma BT325 cell lines. Methods An oligo sequence of shRNA was designed and constructed for GFP gene, and co-transfected with report gene pEGFP-N1 to BT325 cell. After confirming the optimal ratio of transfection, expression of GFP and depressing effect of shRNA on GFP were observed under fluorescence microscope. Results The plasmid carrying GFP gene was cut into 6 kb and 100 bp by restriction endonuclease analysis, which was proved to be the same as original construction. The highest expression of GFP was seen when the ratio of transfection for plasmid DNA and XP-1 was 1︰2.5. The expression of GFP was obviously inhibited by GFP shRNA after 48 h transfection. Conclusions RNA interference exists in BT325 cell line. shRNA could induce specific and stable inhibition of GFP expression, which will provide a new strategy for researching gene function in cancer cell lines.