Epidermolysis Bullosa (EB) is a group of inherited skin blistering diseases which is associated with severely compromised integrity of the skin. One of the most severe form is recessive dystrophic EB (RDEB) caused by loss-of-function mutations in the COL7A1 gene, encoding type VII collagen (COL7). Deficiency of COL7 results in skin fragility, leading to severe recurrent trauma-induced blistering of the skin. Currently, there is no effective therapy or cure for RDEB. Several treatment approaches are ongoing, but the lack of representative skin models of the physiology and structure of the skin have limited such studies to date. Here, induced pluripotent stem cell (iPSC) technology was used in conjunction with genome editing on previously derived iPSCs heterozygous for a mutation in COL7A1 gene (carrier RDEB-iPSC or cRDEB-iPSC) to generate isogenic cRDEB-iPSC lines by complete knock-out of the COL7A1 gene. These can be further used for the generation of 3D human skin organoids and modeling disease in vitro. First, successful reprogramming of the cRDEB-iPSC line was validated in vitro by pluripotency and differentiation capacity assays. CRISPR-Cas9 targeted gene knock-out upon ribonucleoprotein (RNP) transfection of cRDEB-iPSCs using two different RNPs yielded at least one potential knock-out of the COL7A1 gene. However, COL7 expression must be further assessed to confirm complete COL7A1 gene knock-out on the wild-type allele. Nonetheless, the project demonstrated the success in combining the novel iPSCs and CRISPR-Cas9 technologies but revealed the complexity and possible complications associated with their application.