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Recommendations for Successful siRNA Library Screens

type can affect siRNA screening results, as illustrated in the following example. A screen of siRNAs transfected into HeLa cells revealed three genes whose reduction resulted in a cell proliferation defect. siRNAs targeting these three genes as well as nine other genes were re-transfected into HeLa cells as well as into HepG2 cells. The cell proliferation assay confirmed that reducing the expression of the three genes reduced cell proliferation in HeLa cells (Figure 6A). However, siRNAs targeting only one of the three genes reduced cell proliferation in HepG2cells. Instead, two other genes appear to be involved in cell proliferation in HepG2 cells (Figure 6). These results point out the fundamental differences between cell types. Different cells express different genes and feature different cellular pathways. Depending on the assay being used, it is likely that different cell types will reveal different hits. This points to the importance of selecting appropriate cells for the biological function being studied and highlights the opportunity to identify many interesting target genes by using multiple cell types in screening experiments.

Figure 6. Response to Specific siRNA-mediated Gene Inhibition Varies with Cell Type. HeLa (A) and HepG2 (B) cells were transfected in triplicate with three siRNAs targeting 13 different genes or a scrambled siRNA negative control. Cells were counted three days after transfection. Although MAPK13 (brown) inhibition decreases cell proliferation in both cell types, CDK7 (red) and GAPDH (blue) inhibition preferentially decreases cell proliferation in HeLa cells, and MAPK12 (yellow) preferentially decreases cell proliferation in HepG
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