Targeted disruption of the alpha1,3-galactosyltransferase gene in cloned pigs.
Galactose-alpha1,3-galactose (alpha1,3Gal) is the major xenoantigen causing hyperacute rejection in pig-to-human xenotransplantation. Disruption of the gene encoding pig alpha1,3-galactosyltransferase (alpha1,3GT) by homologous recombination is a means to completely remove the alpha1,3Gal epitopes from xenografts. Here we report the disruption of one allele of the pig alpha1,3GT gene in both male and female porcine primary fetal fibroblasts. Targeting was confirmed in 17 colonies by Southern blot analysis, and 7 of them were used for nuclear transfer. Using cells from one colony, we produced six cloned female piglets, of which five were of normal weight and apparently healthy. Southern blot analysis confirmed that these five piglets contain one disrupted pig alpha1,3GT allele.
Dai
Yifan
Y
PPL Therapeutics Inc., 1700 Kraft Drive, Blacksburg, VA 24060, USA. ydai@ppl-therapeutics.com
Vaught
Todd D
TD
Boone
Jeremy
J
Chen
Shu-Hung
SH
Phelps
Carol J
CJ
Ball
Suyapa
S
Monahan
Jeff A
JA
Jobst
Peter M
PM
McCreath
Kenneth J
KJ
Lamborn
Ashley E
AE
Cowell-Lucero
Jamie L
JL
Wells
Kevin D
KD
Colman
Alan
A
Polejaeva
Irina A
IA
Ayares
David L
DL
D000818:Animals; D015139:Blotting, Southern; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D019976:Cloning, Organism; D000939:Epitopes; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D005700:Galactosyltransferases / Q000235:genetics*; D008297:Male; D008957:Models, Genetic; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic; D013552:Swine / Q000235:genetics*; D014162:Transfection
D000939:Epitopes; D005700:Galactosyltransferases; C047006:N-acetyllactosaminide alpha-1,3-galactosyltransferase