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. 2008 Nov 6;36(22):7100–7109. doi: 10.1093/nar/gkn902

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© 2008 The Author(s)

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 1. — Off-target gene silencing assay using reporter plasmids. Structures of (A) psiCHECK-cm and (B) psiCHECK-sm (the luc transcript of which, respectively, presents three tandem repeats of cm and sm in the 3′ UTR, shown with thick arrows). (C) Gene-silencing activities of 26 siRNAs were examined in HeLa cells as a function of siRNA concentration. In most cases, luc mRNA with cm was effectively inactivated. In contrast, inactivation of mRNA with sm varied significantly, depending on the siRNA concentration used for transfection. Sequences of siRNA, cm targets and sm targets are shown in Supplementary Tables S1 and S2. (D) Simultaneous reduction in mRNA amount and luc activity in psiCHECK-transfected cells. Ordinate represents relative luc activity (%) in different psiCHECK-sm and -cm transfected cells. Abscissa represents relative amount of luc mRNA (%) in the corresponding psiCHECK-sm and -cm transfected cells. Results strongly suggest that the reduction of luc activity (protein activity) is due mainly to gene-silencing-dependent degradation of the corresponding mRNA.