Several studies have led to the development of guidelines for the generation of siRNAs which are optimal in terms of efficacy and specificity [12, 16] . This includes the initial definition of the preferable length (19-25 bp) combined with a low G/C content in the range between 36% and 52% and the requirement of symmetric 2 nt overhangs at the 3 -end [16] [17] [18] . Later studies on synthetic siRNA molecules, however, revealed an up to 100-fold higher targeting efficacy in the case of even longer duplexes (25-30 nucleotides) which act as a substrate for Dicer and which therefore allow the direct incorporation of the newly produced siRNAs into the RISC complex [19] . As to be expected, intramolecular foldback structures which can result from internal repeats or palindrome sequences decrease the numbers of functional siRNA molecules with silencing capability [20] . Additional silencing-enhancing criteria include an A in position 3 and a G at position 13 of the sense strand, the absence of a C or G at position 19 and, most importantly, a U in position 10 of the sense strand. Since nucleotides 10-11 represent the site of the RISC-mediated cleavage of the target mRNA, this indicates that RISC is comparable to most other endonucleases in preferentially cleaving 3 of U rather than any other nucleotide [20, 21] . Furthermore, it was shown more generally that the thermodynamic flexibility of the positions 15-19 of the sense strand correlates with the silencing efficacy and that the presence of at least one A/U base pair in this region improves siRNA-mediated silencing efficacy due to a decreased internal stability of its 3 -end [20] .