title: Production of alpha-1,3-galactosyltransferase null pigs by means of nuclear transfer with fibroblasts bearing loss of heterozygosity mutations.
abstract: Hyperacute rejection of porcine organs by old world primate recipients is mediated through preformed antibodies against galactosyl-alpha-1,3-galactose (Galalpha-1,3-Gal) epitopes expressed on the pig cell surface. Previously, we generated inbred miniature swine with a null allele of the alpha-1,3-galactosyltransferase locus (GGTA1) by nuclear transfer (NT) with gene-targeted fibroblasts. To expedite the generation of GGTA1 null pigs, we selected spontaneous null mutant cells from fibroblast cultures of heterozygous animals for use in another round of NT. An unexpectedly high rate of spontaneous loss of GGTA1 function was observed, with the vast majority of null cells resulting from loss of the WT allele. Healthy piglets, hemizygous and homozygous for the gene-targeted allele, were produced by NT by using fibroblasts that had undergone deletional and crossover/gene conversion events, respectively. Aside from loss of Galalpha-1,3-Gal epitopes, there were no obvious phenotypic differences between these null piglets and WT piglets from the same inbred lines. In fact, congenital abnormalities observed in the heterozygous NT animals did not reappear in the serially produced null animals.
authors.affiliation: Immerge Biotherapeutics, Inc., 300 Technology Square, Cambridge, MA 02139, USA.
authors.forename: Donna
authors.initials: D
authors.lastname: Kolber-Simonds
authors.forename: Liangxue
authors.initials: L
authors.lastname: Lai
authors.forename: Steven R
authors.initials: SR
authors.lastname: Watt
authors.forename: Maria
authors.initials: M
authors.lastname: Denaro
authors.forename: Scott
authors.initials: S
authors.lastname: Arn
authors.forename: Monica L
authors.initials: ML
authors.lastname: Augenstein
authors.forename: Jeffery
authors.initials: J
authors.lastname: Betthauser
authors.forename: David B
authors.initials: DB
authors.lastname: Carter
authors.forename: Julia L
authors.initials: JL
authors.lastname: Greenstein
authors.forename: Yanhong
authors.initials: Y
authors.lastname: Hao
authors.forename: Gi-Sun
authors.initials: GS
authors.lastname: Im
authors.forename: Zhonghua
authors.initials: Z
authors.lastname: Liu
authors.forename: Greg D
authors.initials: GD
authors.lastname: Mell
authors.forename: Clifton N
authors.initials: CN
authors.lastname: Murphy
authors.forename: Kwang-Wook
authors.initials: KW
authors.lastname: Park
authors.forename: August
authors.initials: A
authors.lastname: Rieke
authors.forename: David J J
authors.initials: DJ
authors.lastname: Ryan
authors.forename: David H
authors.initials: DH
authors.lastname: Sachs
authors.forename: Erik J
authors.initials: EJ
authors.lastname: Forsberg
authors.forename: Randall S
authors.initials: RS
authors.lastname: Prather
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Hawley
mesh_terms: D000818:Animals; D015139:Blotting, Southern; D002460:Cell Line; D005347:Fibroblasts / Q000648:ultrastructure; D005434:Flow Cytometry; D005700:Galactosyltransferases / Q000235:genetics*; D019656:Loss of Heterozygosity; D053652:Nuclear Transfer Techniques; D010641:Phenotype; D013552:Swine
chemical_list: D005700:Galactosyltransferases