In addition to or as an alternative to siRNA duplexes, shRNA vectors are frequently used for RNAi applications. Vectors expressing shRNA offer an advantage over siRNA duplexes in that they permit a longer-lived inhibition of target genes. As transfection efficiency is one of the two most important factors determining the effectiveness of RNAi, the ability of the Nucleofector™ Technology to transfect a wide range of cells with high efficiencies significantly increases the range of cell types in which plasmid based RNAi systems can be used, such as primary neurons.
Benefits of Nucleofection™ for shRNA-based RNAi applications:
- Up to 90% transfection efficiency for DNA vectors even in suspension cell lines and primary cells
- Non-viral transfection circumvents laborious viral transduction - cost and time effective
- Identical transfection conditions for DNA or RNA - simplifies switch between substrates
HT Nucleofector™ System or 96-well Shuttle™ System enabling shRNA library screening
Knockdown of Fas surface expression on Jurkat cells by Nucleofection™ of a lentiviral anti-Fas shRNA vector. Jurkat E6-1 were transfected by Nucleofection™ with either 0.5 µg shRNA vector targeting Fas or 0.5 µg pmaxGFP™ Vector (negative control) using Cell Line 96-well Nucleofector™ Solution SE and program CL-120. Cells were stained at various time points post Nucleofection™ with an antibody directed against Fas and analyzed by flow cytometry. Staining intensity is normalized to negative control expression of Fas (set to 100% for each time point).