NR ATON
AU Klose,R.; Wünsch,A.; Zakhartchenko,V.; Yang,F.; Wenigerkind,H.; Reichenbach,H.D.; Mitko,K.; Wolf,E.
TI Towards PrPc-deficient cattle: generation of pre-implantation embryos and establishment of pregnancies using PRNP-/- fibroblasts for somatic cell nuclear transfer
QU International Conference - Prion 2005: Between fundamentals and society's needs - 19.10.-21.10.2005, Congress Center Düsseldorf - Poster Session: Human prions, risk of blood products, and therapy HUMAN-37
PT Konferenz-Poster
AB
Cattle lacking PrPc are important for evaluation of the physiological functions of PrPc in bovine and for clarifying the role of PrPc in the pathogenesis of BSE. If PrPc-deficient cattle prove to be resistant to BSE as is expected from studies in Prnp-/- mice, such cattle are very important for animal biotechnology, providing a source of biopharmaceutical products devoid the risk for BSE infection.
Due to the lack of germ-line competent embryonic stem cells in livestock species, nuclear transfer using targeted donor cells is so far the only possibility to perform gene knock-outs in livestock species. Targeted inactivation of the PRNP gene in primary fetal fibroblasts is achieved by homologous recombination using promoter trap vectors in which the entire coding sequence for PrPc is replaced by promoter-less resistance genes (neomycin, hygromycin).
We inactivated the first allele of the PRNP gene with the help of a neomycin-vector. Targeted fibroblasts were used for nuclear transfer cloning to produce PRNP+/- fetuses. We established primary fibroblast cultures of three fetuses and verified the PRNP+/- genotype by Southern blot. The second PRNP allele was inactivated in PRNP+/- cells applying the same strategy but with a promoter-less hygromycin resistance gene. From those, we established pregnancies using PRNP-/- nuclear transfer embryos. Southern blot analysis of fetal fibroblasts derived from five fetuses showed correct targeting of both alleles of the PRNP gene. These PRNP-/- cells are currently being used for nuclear transfer to produce PrPc-deficient offspring. Pregnancies of different ages have been established so far.
IN Die Autoren versuchen, PrPc-knock-out-Rinder zu erzeugen, die mangels Prionprotein-Expression nicht BSE-infiziert werden können und daher garantiert BSE-freies biologisches Material liefern können. Außerdem könnten natürlich solche Rinder nützlich für die Aufklärung der Rollen des normalen Prionproteins sein.
AD R.Klose, A.Wünsch, V.Zakhartchenko, F.Yang, H.Wenigerkind, K.Mitko, E.Wolf, Ludwig-Maximilians-Universität München, Lehrstuhl für Molekulare Tierzucht, Germany; H.D.Reichenbach, Bayerische Landesanstalt für Landwirtschaft, Institut für Tierzucht, Germany
SP englisch
PO Deutschland