Production of a1,3-galactosyltransferase targeted pigs using transcription activator-like effector nuclease-mediated genome editing technology

• Published in: Journal of veterinary science (Suwŏn-si, Korea) pp. 89 - 96
• Main Authors: 강정택, Dae-Kee Kwon, A-Rum Park, Eun-Jin Lee, Yun-Jin Yun, Dal-Young Ji, Kiho Lee, 박광욱
• Format: Journal Article
• Language: English
• Published: 대한수의학회 01.03.2016
• Subjects: 수의학
• ISSN: 1229-845X; 1976-555X
• DOI: 10.4142/jvs.2016.17.1.89

Recent developments in genome editing technology using meganucleases demonstrate an efficient method of producing gene edited pigs. In the present study, we examined the effectiveness of the transcription activator-like effector nuclease (TALEN) system in generating specific mutations on the pig genome. Specific TALEN was designed to induce a double-strand break on exon 9 of the porcine a1,3-galactosyltransferase (GGTA1) gene as it is the main cause of hyperacute rejection after xenotransplantation. Human decay-accelerating factor (hDAF) gene, which can produce a complement inhibitor to protect cells from complement attack after xenotransplantation, was also integrated into the genome simultaneously. Plasmids coding for the TALEN pair and hDAF gene were transfected into porcine cells by electroporation to disrupt the porcine GGTA1 gene and express hDAF. The transfected cells were then sorted using a biotin-labeled IB4 lectin attached to magnetic beads to obtain GGTA1 deficient cells. As a result, we established GGTA1 knockout (KO) cell lines with biallelic modification (35.0%) and GGTA1 KO cell lines expressing hDAF (13.0%). When these cells were used for somatic cell nuclear transfer, we successfully obtained live GGTA1 KO pigs expressing hDAF. Our results demonstrate that TALEN-mediated genome editing is efficient and can be successfully used to generate gene edited pigs. KCI Citation Count: 13