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This analysis determined that the optimal cell concentration necessary to induce maximum interference was 2000 cells per well (Figure 1). Data clearly show that the optimal number of HeLa cells plated in this experiment was 2,000 cells (Figure 1). At this plating density, cells transfected with the siRNA targeting GAPDH expressed only 30% the GAPDH levels of those transfected with the negative control siRNA.

Figure 1. Optimization of siRNA Transfection Using Cells-to-cDNA II Automated Protocol. The Y axis shows GAPDH expression after transfection with GAPDH siRNA as a percent of expression after transfection with the negative control siRNA. A standard curve for GAPDH expression was used to normalize the data. An ABI 7900 was used to perform realtime PCR.

Ambion's Silencer siRNA products are described in more detail in, "Cost Effective High Potency siRNAs". For more information about the Cells-to-cDNA II automated protocol, see "Automation of One-Step RT-PCR Using Ambion's Cells-to-cDNA II Kit and the MultiPROBE II HT Liquid Handling System" or contact our technical service department by email at http://www.ambion.com/contact/techserv_email.html or phone at (800) 888-8804 option 2. Our customers frequently develop new and interesting adaptations of our Cells-to-cDNA technology. Do you have an unusual Cells-to-cDNA story? If so we would like to know!

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