Sequence-specific silencing effect of the shRNA-intron. (A) The firefly luciferase sequences in the pGL3-Control vector that correspond to the targeting regions of 155-173 and 851-875 have a slightly different sequence composition than those in the psiCHECK-2 vector. The sequences from the pGL3-Control plasmid were synthesized and cloned downstream from the stop codon of the Renilla luciferase gene in the psiCHECK-2 vector. (B) The sequence-specific repression by shRNA interrupted by an intron was analyzed by transient cotransfection into Cos cells with the GL155 or GL851 vector. The empty pRNA-CMV3.1-Neo vector was used as a control. Luciferase activity was assessed using the Dual-luciferase Reporter Assay System, in which Renilla luciferase activity was normalized to firefly luciferase activity, and the data are shown as the mean ± S.D. of the percentage of the value for the empty psiCHECK-2 vector.