Genetically modified pigs produced with a nonviral episomal vector.
Genetic modification of cells and animals is an invaluable tool for biotechnology and biomedicine. Currently, integrating vectors are used for this purpose. These vectors, however, may lead to insertional mutagenesis and variable transgene expression and can undergo silencing. Scaffold/matrix attachment region-based vectors are nonviral expression systems that replicate autonomously in mammalian cells, thereby making possible safe and reliable genetic modification of higher eukaryotic cells and organisms. In this study, genetically modified pig fetuses were produced with the scaffold/matrix attachment region-based vector pEPI, delivered to embryos by the sperm-mediated gene transfer method. The pEPI vector was detected in 12 of 18 fetuses in the different tissues analyzed and was shown to be retained as an episome. The reporter gene encoded by the pEPI vector was expressed in 9 of 12 genetically modified fetuses. In positive animals, all tissues analyzed expressed the reporter gene; moreover in these tissues, the positive cells were on the average 79%. The high percentage of EGFP-expressing cells and the absence of mosaicism have important implications for biotechnological and biomedical applications. These results are an important step forward in animal transgenesis and can provide the basis for the future development of germ-line gene therapy.
Manzini
Stefano
S
Department of Surgical Sciences, University of Milano-Bicocca, 20052 Milan,Italy.
Vargiolu
Alessia
A
Stehle
Isa M
IM
Bacci
Maria Laura
ML
Cerrito
Maria Grazia
MG
Giovannoni
Roberto
R
Zannoni
Augusta
A
Bianco
Maria Rosaria
MR
Forni
Monica
M
Donini
Pierluigi
P
Papa
Michele
M
Lipps
Hans J
HJ
Lavitrano
Marialuisa
M
D000818:Animals; D030801:Animals, Genetically Modified; D005333:Fetus / Q000033:anatomy & histology / Q000502:physiology; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics* / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D010957:Plasmids / Q000235:genetics* / Q000378:metabolism; D013094:Spermatozoa / Q000502:physiology; D013552:Swine; D014018:Tissue Distribution; D019076:Transgenes
D049452:Green Fluorescent Proteins