Because knockdown of gene expression in some cell types may not correlate 100% with uptake of fluorescently labeled siRNA, the siPORT siRNA Electroporation Kit also includes a positive control siRNA that targets GAPDH. The unlabeled GAPDH siRNA control can be used in conjunction with the unlabeled negative control siRNA (also included) to refine electroporation conditions by monitoring knockdown of GAPDH mRNA levels. The negative control siRNA is important not only for optimization experiments, but also for any siRNA experiment to confirm the absence of nonspecific effects due to transfection.
The siPORT siRNA Electroporation Kit has been tested with two commonly used single-cuvette electro-pulse generators: Gene Pulser Xcell (Bio-Rad) and ECM 830 (BTX). Their performance was similar when the same electroporation parameters were used.
Effective Silencing in Primary Cells
Figure 2 shows gene silencing induced by several different
siRNAs, all delivered into hMSC and NHDF-neo cells (both are primary cells:
human mesenchymal stem cells, and normal human dermal fibroblasts-neonatal)
by electroporation using siPORT siRNA Electroporation Buffer. A critical
parameter in the success of any RNAi experiment is the potency and specificity
of siRNA sequences. To ensure the success of these gene-silencing experiments