Extended Data Fig. 2: Optimization of canine μUtro transgene expression.
From: Non-immunogenic utrophin gene therapy for the treatment of muscular dystrophy animal models
a, Schematic representation of μUtro based on CnN3D renderings of calponin homology domain (green), four spectrin-like repeats (yellow/blue) and combined WW-EF hand-ZZ domain (red). Disordered domains (‘hinges’) 1, 2 and 4 not depicted. a′, Western blot analysis and quantification of μUtro expression in HEK 293 cells using eight distinct plasmid vectors. a′, Western blot analysis and quantification of μUtro expression one week post intramuscular injections in the tibialis anterior (TA) followed by electroporation of six distinct μUtro containing vectors, as labeled (UTO, μ-Utro cDNA; opt, optimized; SP, synthetic promoter C5-1253; loading control, α-actin; CMV, cytomegalovirus promoter). b,c, Immunofluorescence staining against utrophin N terminus (Utro_N) and laminin in mdx mice injected with either Anc80-μUtro or PBS. Muscles collected include diaphragm, heart (b), quadriceps (quad), TA and gastrocnemius (GM) (c). d, Western blot analysis of μUtro expression in tissue samples, with vinculin serving as a loading control. e, Western blot analysis comparing μUtro expression in the TA and GM after systemic deliver using an AAV9 or Anc80 vector. Vinculin serving as a loading control. (See Source Data for full uncropped gel images.) All experiments were repeated independently at least two or more times with similar results. (See Source Data Extended Data Fig. 2).
