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. 2005 Feb 1;33(2):796–804. doi: 10.1093/nar/gki220

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© The Author 2005. Published by Oxford University Press. All rights reserved

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Stability of the S hairpin determines the level of resistance. (A) Mutant pGL3-Nef reporter constructs with nucleotide substitutions upstream of the siRNA-Nef target sequence were constructed (mutations at positions −7 and/or −26 are in bold and underlined). The stability (ΔG) of the S and R structures was predicted with the Mfold program. The ΔΔG, providing a measure of the S and R structure equilibrium, was calculated (ΔΔG= ΔG^S − ΔG^R). (B) Luciferase expression after transfection of the reporter constructs with increasing amounts of pBS-siRNA-Nef. The level of expression in the absence of siRNA-Nef was set at 100% for each reporter construct. (C) The ΔΔG of the structures is plotted against the level of resistance as observed in Figure 6B with 5 ng of pBS-siRNA-Nef.