Usher syndrome is the most common form of inherited deaf-blindness. Mutations in MYO7A are the predominant cause for USH1B, the most severe subtype of Usher syndrome. There is currently no therapy that can halt or mitigate retinal degeneration in USH1B patients. As the MYO7A coding sequence (6.7 kb) exceeds the packaging capacity of adeno-associated-viral (AAV) vectors, alternative strategies including dual AAV approaches are required for gene supplementation. Here, we used a novel dual AAV vector strategy based on reconstitution via mRNA trans-splicing (REVeRT) to assemble MYO7A in mice, pigs and human retinal organoids. Upon subretinal injection, we show that our dual REVeRT AAV vectors lead to high MYO7A reconstitution efficiency (up to 56%) at protein level in the mouse and pig retina. A side-by-side comparison of REVeRT AAVs to a commonly used dual AAV approach based on reconstitution at the genomic level shows a 3-fold increase in MYO7A expression at transcript and 38% higher expression at protein level. The transgenic MYO7A protein was found to be mainly localized to the RPE and photoreceptor cells of mice, pigs and human retinal organoids. Our results provide compelling evidence that REVeRT leads to high expression of the full-length MYO7A protein in different species. A second strategy aiming at dCas9-VPR-mediated transactivation of MYO7B as a functional equivalent counterpart to MYO7A also shows promising data in in-vitro and in-vivo experiments. Further experiments are currently under investigation. After validation of our dual REVeRT-AAV vectors in Myo7a-deficient animals, we plan to initiate clinical trials in USH1B patients.