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Five Ways to Produce siRNAs

gents necessary to prepare 20 different SECs with either the human H1, human U6, or mouse U6 promoter.

pSEC vectors, which are pre-linearized with the same restriction enzymes and are designed especially for subcloning SECs, are available separately. These ready-to-ligate vectors include a hygromycin, neomycin or puromycin resistance gene to permit selection in mammalian cells.

Figure 4. Variable Reduction in Target Gene Expression Using SECs with Different Promoters. siRNA Expression Cassettes featuring the mouse U6 (Mo-U6), human U6 (Hu-U6), and human H1 (Hu-H1) promoters and encoding a c-fos-specific hairpin siRNA were transfected into HeLa cells. 72 hours post-transfection, the cells were assessed using nuclear staining with DAPI and immunofluorescence using a c-FOS antibody. Non-transfected cells (NT) as well as cells transfected with an SEC expressing a negative control siRNA (scramble) demonstrate wild-type levels of c-FOS. The relative level of reduction in gene expression was quantified and is provided in the bar graph.


siRNA and siRNA Expression Vector Delivery
Once siRNAs or siRNA expression vectors are obtained, they must be delivered to cells. Articles Optimizing Chemical Transfection and Electroporation of siRNAs and Efficient Delivery of siRNAs to Human Primary Cell
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