Production of biallelic CMP-Neu5Ac hydroxylase knock-out pigs

• DOI: 10.1038/srep01981
• Publication Date: 2013-06-13T09:08:28Z
• Authors: Deug-Nam Kwon; Kiho Lee; Man-Jong Kang; Yun-Jung Choi; Chankyu Park; Jeffrey J. Whyte; Alana N. Brown; Jae-Hwan Kim; Melissa Samuel; Jiude Mao; Kwang-Wook Park; Clifton N. Murphy; Randall S. Prather; Jin-Hoi Kim
• Keywords: Male; 0301 basic medicine; Binding Sites; DNA End-Joining Repair; Base Sequence; Genetic Vectors; Homozygote; Karyotype; Molecular Sequence Data; Gene Expression; Fibroblasts; Article; Mixed Function Oxygenases; Animals, Genetically Modified; Gene Knockout Techniques; 03 medical and health sciences; Genetic Loci; Gene Order; Gene Targeting; Animals; Female; Amino Acid Sequence; Homologous Recombination

After the knock-out (KO) of α1,3 galactosyltransfease (Gal-T), the Hanganutziu-Deicher antigen became a major antigen of the "non-Gal antigen" that is implicated in subsequent xenograft rejection. For deletion of non-Gal antigen, we successfully produced zinc finger nuclease (ZFN)-mediated monoallelic/biallelic male and female CMP-N-acetylneuraminic acid hydroxylase (CMAH) KO miniature pigs: the efficiency of the gene targeting (41.7%) was higher when donor DNA was used with the ZFN than those of ZFN alone (9.1%). Monoallelic KO pigs had no integration of exogenous DNA into their genome, indicating that this technique would provide a new avenue to reduce the risk of antibiotics resistance when organs from genetically modified pigs are transplanted into patients. Until now, both monoallelic and biallelic CMAH KO pigs are healthy and show no sign of abnormality and off-target mutations. Therefore, these CMAH null pigs on the Gal-T KO background could serve as an important model for the xenotransplantation.