Objective The shortage of donors is a major problem in organ transplantation, and the generation ofhumanized organs is an effective method to solve this problem. A lack of fumarylacetoacetate hydrolase (FAH) causeshereditary tyrosinemia type I (HTI), which leads to hepatocyte apoptosis and liver damage. Genetic editing of pigs (Susscrofa), establishment of FAH knockout Bama minipigs, combined with human stem cells will facilitate the production ofhumanized stem cell liver. In this study, FAH gene knockout Bama minipigs were generated by CRISPR/ Cas9 based onalpha-1,3-galactosyltransferase (GGTA1) gene knockout pigs,and their health and breeding status were analyzed. Methods Single guide RNA (sgRNAs) were designed to target the porcine FAH gene to construct a pX330 vector that wastransfected into Bama minipig ear fibroblasts. The efficiency of this method was verified. After identification, positive cellswere selected as nuclear donors, and FAH gene knockout Bama minipigs were generated by somatic cell nucleartransplantation technology. The genomic DNA of piglets was extracted and the mutant type was obtained by PCR sequencing. FAH^{+/ -} Bama minipig blood was collected at the age of 5 months, and its blood biochemical and blood routine indexes weretested. Histological changes of livers were analyzed. When FAH^{+/ -} Bama minipigs were sexually mature, they were matedwith wild type (WT) sows, and their breeding ability was evaluated by litter size. Results FAH^{+/ -} and FAH^{-/ -} Bamaminipigs were obtained. Western blotting showed that FAH^{+/ -} Bama minipigs had decreased FAH expression in the liver andkidney. There were no significant differences in the physical and chemical indexes of blood from the FAH^{+/ -} Bama minipigsor WT minipigs. Histological examination revealed vacuolar-like changes in the FAH^{+/ -} pig liver tissues. The litter size ofwild sows bred with FAH^{+/ -} boars was normal. FAH^{+/ -} pigs had normal health status and breeding ability. Conclusions FAH single allele knockout pigs are generated in this study, with normal health and reproductive abilities, which provides a good basis for future studies of the production of double-allele knockout pigs and humanized liver.