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. 2009 Aug 11;17(10):1712–1723. doi: 10.1038/mt.2009.176

Figure 3.

Figure 3

RNAi activity of lhRNAs of different hairpin stem length. (a) Design of two sets of lhRNAs targeting a consecutive sequence of the HIV-1 nef and r/t gene of 66, 88, and 92 bp in length (lhNef and lhR/T). The shRNA targeting nef and r/t is placed at the base of the hairpin and is marked in green or purple, respectively. Mutations were introduced in the passenger strand of the hairpin (indicated in red) to create G-U bp in the hairpin stem (b) HEK 293T cells were co-transfected with the lhRNA constructs, the corresponding Luc-nef or Luc-r/t reporter and pRL. Two days post-transfection luciferase activities were measured and used to calculate the relative luciferase expression (firefly/renilla ratio). Luciferase expression in the presence of pBS was set at 100%. A scrambled hairpin (SCR) was used as negative control and the original shRNA, shNef, or shR/T, as positive control. Bars represent the average values from five independent transfections and error bars show the SD. bp, base pair; HEK, human embryonic kidney; lhRNA, long hairpin RNA; RNAi, RNA interference; shRNA, short hairpin RNA.