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https://hdl.handle.net/2440/8109
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Type: | Journal article |
Title: | An efficient method for production α(1,3)-galactosyltransferase gene knockout pigs |
Other Titles: | An efficient method for production alpha(1,3)-galactosyltransferase gene knockout pigs |
Author: | Harrison, S. Boquest, A. Grupen, C. Faast, R. Guildolin, A. Giannakis, C. Crocker, L. McIlfatrick, S. Ashman, R. Wengle, J. Lyons, I. Tolstoshev, P. Cowan, P. Robins, A. O'Connell, P. d'Apice, A. Nottle, M. |
Citation: | Cellular Reprogramming, 2004; 6(4):327-331 |
Publisher: | Mary Ann Liebert, Inc. Publishers |
Issue Date: | 2004 |
ISSN: | 1536-2302 1557-7457 |
Statement of Responsibility: | Sharon Harrison, Andrew Boquest, Christopher Grupen, Renate Faast, Angelo Guildolin, Christopher Giannakis, Lesley Crocker, Stephen McIlfatrick, Rodney Ashman, James Wengle, Ian Lyons, Paul Tolstoshev, Peter Cowan, Allan Robins, Philip O'Connell, Anthony J.F. D'Apice and Mark Nottle |
Abstract: | We have reported relatively efficient methods for somatic cell nuclear transfer and for knocking out the alpha(1,3)-galactosyltransferase (alpha1,3-GT) gene in porcine fetal fibroblasts using a nonisogenic promoterless construct approach. Here we report the production of alpha1,3-GT gene knockout pigs using these procedures. Seven alpha1,3-GT gene knockout cell clones were identified by long-range PCR from 108 neomycin resistant (neo(R)) colonies, giving a 6.5% targeting efficiency. Three cell clones were used for nuclear transfer. Nuclear transfer was performed using a fusion before activation protocol using in vitro-matured adult oocytes. Between 51 and 110 fused couplets were transferred to 10 recipients synchronized 1 day behind the embryos. Parturition was induced on day 115, and piglets were delivered by caesarean section. Four recipients gave birth to a total of 18 live piglets. All pigs were female, and all three clones resulted in the birth of live pigs. alpha1,3-GT gene knockout pigs were identified by long-range PCR and confirmed by Southern blot analysis. The efficiency (embryos transferred/piglets born) of our cloning protocol was 1.9% for all transfers and 4.6% for animals that gave birth. |
Keywords: | Oocytes Fibroblasts Animals Animals, Genetically Modified Swine Galactosyltransferases Gene Deletion Pregnancy Female Nuclear Transfer Techniques |
DOI: | 10.1089/clo.2004.6.327 |
Published version: | http://dx.doi.org/10.1089/clo.2004.6.327 |
Appears in Collections: | Aurora harvest 4 Obstetrics and Gynaecology publications |
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