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[SANET-MG] genetic modification to make unmadable cows



RNA interference technology is new and its stability after genetic modification is unpredictable. It is probably be unwise to run out and buy stock in an RNA interference company.
Published online before print March 27, 2006
Proc. Natl. Acad. Sci. USA, 10.1073/pnas.0600813103
Agricultural Sciences
Suppression of prion protein in livestock by RNA interference

Michael C. Golding *, Charles R. Long , Michelle A. Carmell *, Gregory J. Hannon *, and Mark E. Westhusin *Watson School of Biological Sciences, Cold Spring Harbor Laboratory, Howard Hughes Medical Institute, 1 Bungtown Road, Cold Spring Harbor, NY 11724; and Department of Veterinary Physiology, College of Veterinary Medicine, Texas A & M University, College Station, TX 77843

Given the difficulty of applying gene knockout technology to species other than mice, we decided to explore the utility of RNA interference (RNAi) in silencing the expression of genes in livestock. Short hairpin RNAs (shRNAs) were designed and screened for their ability to suppress the expression of caprine and bovine prion protein (PrP). Lentiviral vectors were used to deliver a transgene expressing GFP and an shRNA targeting PrP into goat fibroblasts. These cells were then used for nuclear transplantation to produce a cloned goat fetus, which was surgically recovered at 81 days of gestation and compared with an age-matched control derived by natural mating. All tissues examined in the cloned fetus expressed GFP, and PCR analysis confirmed the presence of the transgene encoding the PrP shRNA. Most relevant, Western blot analysis performed on brain tissues comparing the transgenic fetus with control demonstrated a significant (>90%) decrease in PrP expression levels. To confirm that similar methodologies could be applied to the bovine, recombinant virus was injected into the perivitelline space of bovine ova. After in vitro fertilization and culture, 76% of the blastocysts exhibited GFP expression, indicative that they expressed shRNAs targeting PrP. Our results provide strong evidence that the approach described here will be useful in producing transgenic livestock conferring potential disease resistance and provide an effective strategy for suppressing gene expression in a variety of large-animal models.--

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