Figure 2.

Evaluation of knockdown efficiency between first, second, and third generation vector-based RNAi triggers. Left, knockdown efficiency of ff-luc relative to Rr-luc within distinct hairpin expression contexts. 10 distinct targeting sequences were evaluated with hairpins expressed from a human U6 promoter (Pol-III) or CMV (Pol-II) promoter, all of which were co-transfected in a 1:30 w/w ratio to the target expression plasmid. Evaluated were the first generation shRNA system, TRC, the hybrid first and second generation system, BD, the second generation amiRNA, miR-30, and distinct third generation systems, miR-E and miR-3G. Right, as left, with 8 unique sequences targeting Rr-luc. Knockdown was normalized to firefly luciferase. All knockdown was measured ∼40 hours post-transfection into HEK293T cells. Shown is a representative experiment (N = 2 ) performed in technical triplicate. Each point represents an average of the technical triplicates, and bars represent the mean for individual data sets.