RNAinterference (RNAi) has developed within a short time from an area of basic research occupied by a few experts to a widely used technical tool for reverse genetics, which is expected to have a broad utility not only in research, but also in medical and diagnostic applications. Despite its widespread use, the application of RNAi is often hampered because it difference of only a few nuclectides in the sequence of the target RNA can change the efficiency of a small interfering RNA (siRNA) from high to zero, and publicly available design tools for siRNAs are not yet perfect. We therefore developed and compared RNAi test systems based oil different promoters, reporters, and target sequences. Here, we show that fluorescence-based test systems have obvious disadvantages compared to luciferase-based test systems and that some combinations of promoter, reporter, and target sequences, although Currently in use, are not well suited for testing RNAi effects. (c) 2005 Elsevier Inc. All rights reserved.