Analysis of transfection characteristic of two different fluorescently labeled siRNA targeting PIK3CA gene in vitro

Abstract

Objective To compare transfection properties of two different fluorescently labeled PIK3CA siR-NA and to screen out PIK3CA siRNA with high transfection efficiency and strong anti-quenching ability.Methods Two different fluorescently labeled PIK3CA siRNA was transfected into gastric cancer cell BGC-823 by Lipofectamine 2000.The distribution and quenching of fluorescence were observed by inverted fluorescence microscope.Image J software was used to analyze their transfection efficiency.Real-time quantitative PCR was performed to detect the effect of different fluorescently labeled PIK3CA siRNA on PIK3CA mRNA expression.Results The transfection efficiency and anti-quenching ability of two different fluorescently labeled PIK3CA siRNA were different under the same transfection conditions.The transfection efficiency showed no significant differences between Cy3 or FAM labeled PIK3CA siRNA and negative control siRNA(P ＞ 0.05),but the transfection efficiency of Cy3 labeled PIK3CA siRNA and negative control siRNA was significantly higher than the FAM labeled (P ＜ 0.05).Inhibitory efficacy of target mRNA expression induced by Cy3 labeled PIK3CA siRNA was significantly higher than that of FAM labeled PIK3CA siRNA.Conclusion Cy3 labeled PIK3CA siRNA could act as a good tracer and provide an important evidence for further construction of Cy3 labeled PIK3CA siRNA nanoparticle. Key words: PIK3CA gene; Fluorescently labeled small interfering RNA; Transfection