Human Factor IX Transgenic Sheep Produced by Transfer of Nuclei from Transfected Fetal Fibroblasts

Abstract

Ovine primary fetal fibroblasts were cotransfected with a neomycin resistance marker gene (neo) and a human coagulation factor IX genomic construct designed for expression of the encoded protein in sheep milk. Two cloned transfectants and a population of neomycin (G418)-resistant cells were used as donors for nuclear transfer to enucleated oocytes. Six transgenic lambs were liveborn: Three produced from cloned cells contained factor IX and neo transgenes, whereas three produced from the uncloned population contained the marker gene only. Somatic cells can therefore be subjected to genetic manipulation in vitro and produce viable animals by nuclear transfer. Production of transgenic sheep by nuclear transfer requires fewer than half the animals needed for pronuclear microinjection.

Cassette DNA Fragment for Selection and Sexing of Preimplantation Bovine Transgenic Embryos.

“…Although the recently developed cell-mediated method for producing transgenic livestock by nuclear transfer from stably transfected somatic cells to enucleated unfertilized egg will become a revolutionary technology [18,19], microinjection of DNA into the pronuclei of fertilized embryos is one of the practical and reliable methods for the production of transgenic livestock. To cut down the cost for making transgenic cattles, reducing the number of recipient for an implantation of the microinjected embryo is very important.…”

Evaluation of laser-assisted lentiviral transgenesis in bovine

“…High costs and low efficiency of the available methods to create well expressing transgenic livestock could not be overcome sufficiently, although the importance and high value of such animals is emphasized enthusiastically by many authors (Wall et al, 1985;Eyestone et al, 1998;Piedrahita et al, 1999;Wall et al, 1992;Wall et al, 1997;Clark, 1998;Brem and Brening, 1985;Chan, 1999;Clark and Whitelaw, 2003;Colman, 1996;Denning and Priddle, 2003;Donovan et al, 2005;Eyestone, 1994;Golovan et al, 2001;Grosse-Hovest et al, 2004;Hammer et al, 1985;Haskins et al, 2002;Hofmann et al, 2003;Hofmann et al, 2004;Hyttinen et al, 1994;Melo et al, 2005;Muller and Brem, 1998;Petters, 1994;Schnieke et al, 1997;Seamark, 1994;Wolf et al, 2000) The success of pronuclear microinjection or cloning is evident in the generation of transgenic cattle but its limitations have hindered progress of bovine transgenics (Hodges and Stice, 2003;Maga, 2005).…”

Transgenic bovine as bioreactors: Challenges and perspectives

“…44 After that time, numerous transgenic animals were produced using SCNT for many scientific proposes, including heterologous protein production. SCNT is the major means for producing transgenic large animals, as 90 to 100% of newborns carry the transgenes.…”