The activity of siRNAs targeting SNP sites in the SCA3 model. (A) The target sites and nucleotide sequences of siRNAs directed at rs12895357 SNP in the ATXN3 gene. The nucleotide in the antisense strand of siRNA that directly targets the SNP site is marked in bold. The mismatches introduced into siRNA duplex are in gray boxes. RNA is in lowercase; DNA is in uppercase. (B) The RT-PCR analysis of ATXN3 expression at the transcript level in the SCA3 fibroblasts (GM06153) at 72 h after transfection with 50 nM siRNAs. (C) The western blot analysis of the ataxin-3 levels for the experiment described in (B). (D) The western blot analysis of the ataxin-3 levels in the SCA3 fibroblasts at 72 h after transfection with 2, 10, 50 or 200 nM siRNA G9. (E) The western blot analysis as in (D) for siRNA G10. C - the reference bar indicates the ATXN3 expression level in the cells transfected with control siRNA. In the graphs, the signal intensities were normalized to GAPDH mRNA or protein levels and compared using a one-sample t-test. The error bars represent the standard deviations. The P-value is indicated by asterisks (* p < 0.001; ** 0.001 < p < 0.05).
Fiszer et al. BMC Molecular Biology 2012 13:6 doi:10.1186/1471-2199-13-6