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. 2013 Oct 15;21(12):2148–2159. doi: 10.1038/mt.2013.211

Figure 2.

Figure 2

shRNA constructs show efficient reduction of human SOD1 protein in vitro and in vivo. (a) Sequence alignments between human and mouse SOD1 for the regions targeted by the four different shRNA constructs tested. (b) shRNA sequences were cloned into an H1 expression construct and transiently transfected into 293 cells. Lysates were collected 72 hours posttransfection and analyzed by western blot. (c) Quantification of in vitro suppression of human SOD1 from three separate transient transfections showed >50% reduction in SOD1. (d) shRNA 130 was packaged into AAV9 and injected into SOD1G93A mice at either P1 or P21. Spinal cords (n = 3 per time point) were harvested 3 weeks postinjection and analyzed by western blot for human SOD1 protein levels. (e) Quantification of in vivo suppression of human SOD1 within the spinal cord of ALS mice. P1- and P21-injected spinal cords showed 60 and 45% reductions in mutant SOD1 protein. GAPDH, glyceraldehyde 3 phosphate dehydrogenase; hSOD1, human superoxide dismutase 1; mSOD1, mouse superoxide dismutase 1; P1, postnatal day 1; P21, postnatal day 21.