title: Efficient transgenesis in farm animals by lentiviral vectors.
abstract: Microinjection of DNA is now the most widespread method for generating transgenic animals, but transgenesis rates achieved this way in higher mammals are extremely low. To address this longstanding problem, we used lentiviral vectors carrying a ubiquitously active promoter (phosphoglycerate kinase, LV-PGK) to deliver transgenes to porcine embryos. Of the 46 piglets born, 32 (70%) carried the transgene DNA and 30 (94%) of these pigs expressed the transgene (green fluorescent protein, GFP). Direct fluorescence imaging and immunohistochemistry showed that GFP was expressed in all tissues of LV-PGK transgenic pigs, including germ cells. Importantly, the transgene was transmitted through the germ-line. Tissue-specific transgene expression was achieved by infecting porcine embryos with lentiviral vectors containing the human keratin K14 promoter (LV-K14). LV-K14 transgenic animals expressed GFP specifically in basal keratinocytes of the skin. Finally, infection of bovine oocytes after and before in vitro fertilization with LV-PGK resulted in transgene expression in 45% and 92% of the infected embryos, respectively.
authors.affiliation: Department of Pharmacy, Institute for Pharmacology, Center for Drug Research, Butenandtstrasse 5 (C), Ludwig-Maximilians University, 81377 Munich, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Hofmann
authors.forename: Barbara
authors.initials: B
authors.lastname: Kessler
authors.forename: Sonja
authors.initials: S
authors.lastname: Ewerling
authors.forename: Myriam
authors.initials: M
authors.lastname: Weppert
authors.forename: Barbara
authors.initials: B
authors.lastname: Vogg
authors.forename: Harald
authors.initials: H
authors.lastname: Ludwig
authors.forename: Miodrag
authors.initials: M
authors.lastname: Stojkovic
authors.forename: Marc
authors.initials: M
authors.lastname: Boelhauve
authors.forename: Gottfried
authors.initials: G
authors.lastname: Brem
authors.forename: Eckhard
authors.initials: E
authors.lastname: Wolf
authors.forename: Alexander
authors.initials: A
authors.lastname: Pfeifer
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D002417:Cattle; D004622:Embryo, Mammalian / Q000378:metabolism; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors; D007150:Immunohistochemistry; D016086:Lentivirus; D008856:Microscopy, Fluorescence; D009865:Oocytes / Q000378:metabolism; D013552:Swine / Q000235:genetics*