Xenotransplantation is a solution to alleviate the shortage of human donors for patients in waiting list for organ transplantation. Genetic pig engineering is required to generate suitable transgenic pigs as organ donors together with the development of new clinical immunosuppression protocols. In line with the current state of the art, we modified available αGAL-Neu5Gc-SDA KO pig lines targeting 6 human transgenes into the porcine GGTA1 gene, combining the CRISPR/Cas9 and Cre/lox-RMCE. To achieve this result, 3 different RMCE plasmids were created for targeting the GGTA1 gene by sequential insertion of: a) GGTA1-landing pad (lox2272-5171); b) 1^st RMCE cassette: hCD55/CD46/CD59/p-hTM (lox2272-P-5171); c) 2^nd RMCE cassette: hCD47/HMOX1 (lox2272-P). In a single experiment, primary fibroblasts (male; GGTA1/CMAH/B4GalNT2-KO) were transfected (Nucleofector-Lonza; W024) at the same time with the 3 different RMCE plasmids in combination with: Cas9 protein;1 sgRNAs (GGTA1) and Cre recombinase mRNA. Five days post transfection (dpt) cells were sorted using magnetic beads (CELLection, PanMouse IgG) and plated (⍉ =150 mm dish; 150 cells/dish) for clonal selection. Growing colonies (⍉≅5 mm; 13 dpt) were picked up, splitted and expanded for cryopreservation and molecular analysis. Lysed colonies were used for sequential PCR-screenings to confirm: a) the integration of p-hTM, hCD55/CD46/CD59, hCD47/HMOX1 (6TG); b) the GGTA1 targeting events (left and right side); c) 2^nd RMCE event; d) the absence of Prokaryotic backbone integration. Only transgenic colonies that resulted positive for the 2^nd RMCE event were selected for the phenotypic characterization (ICC and WB) to identify ones to use as nuclear donor for the SCNT experiments. Starting from 165 selected colonies, 135 were lysed and 45 resulted transgenic for all the six transgenes. GGTA1-targeting events were detected in 10 transgenic colonies; 7 resulted positive for the Cre-mediated insertion of the 2^nd RMCE cassette, 3 resulted negative for the prokaryotic backbone integration and their phenotype was characterized by WB and ICC. Final WB and ICC analyses confirmed the ubiquitous expression of hCD55/CD46/CD59/CD47/HMOX1(5TGexpr) in 6 colonies except for p-hTM that is driven by a tissue specific promoter. Based on the molecular and phenotypical findings two colonies (6TG; GGTA1-targeted; 2^nd RMCE positive; 5TGexpr) were selected for a SCNT experiment and the resulting embryos were transferred in a synchronized recipient sow, that was pregnant at 30 days. We demonstrated that transfecting a modular Cre/lox-RMCE system combined with CRISPR/Cas9 it is possible to obtain a targeted integration of 6 human transgenes in a single experiment however random integrations of the transfected cassettes needs to be addressed. This modular platform will permit us to perform new RMCE experiments to quickly integrate additional transgenes in the same locus.