Production of transgenic sheep: Microinjection of the bovine P77 gene construct in Chios sheep breed zygotes

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Pubblicato: Jan 31, 2018
DOI: https://doi.org/10.12681/jhvms.15707
E. VAINAS (Ε. ΒΑΪΝΑΣ)
National Agricultural Research Foundation (N.AG.RE.F), Veterinary Research Institute, Thessaloniki
U. BESENFELDER
Institut fuer Tierzucht und Genetik, vet. M. Un. Wien
V. CHRISTODOULOU (B. ΧΡΙΣΤΟΔΟΥΛΟΥ)
Ν. AG. RE.F, Animal Research Institute, Paralimni Giannitsa
B. KUEHHOLZER
LBI fuer zyto-, immuno-und molekulargenetische Forschung, Wien
G. C. AMIRIDIS (Γ.Σ. ΑΜΟΙΡΙΔΗΣ)
Dept. of Obstetrics and Reproduction, Faculty of Vetertinary Medicine, University of Thessaly
S. BELIBASAKI (Σ. ΜΠΕΛΙΜΠΑΣΑΚΗ)
National Agricultural Research Foundation (N.AG.RE.F), Veterinary Research Institute, Thessaloniki
G. BREM
Institut fuer Tierzucht und Genetik, vet. M. Un. Wien
Abstract

The feasibility of integration of the bovine P77 construct into zygotes and two cell embryos of Chios sheep breed was examined. The P77 gene construct comprised bovine as l casein promoter regulatory sequence fused to the fragment between exons 2 and 9 of the bovine chymosin gene. One of the two pronuclei of the zygote or a nucleus of a blastomere of a two cell embryo was microinjected with 1 pi of DNA solution containing approximately 1000 copies of the gene construct. In total 193 zygotes and 87 two cell embryos collected from 49 donor ewes were used. After 1 - 5 h culture microinjected zygotes (n=175) and two cell embryos (n=76) were transferred by a laparoscopic technique to the oviducts of 68 recipients. 22 recipients (32.4%) gave birth to 29 lambs which correspond to 11.6% of the total number of zygotes - embryos transferred. PCR analysis performed in skin samples of these lambs revealed that in one ewe lamb the P77 construct has been integrated in its genome. The growth and puberty of this lamb was physiological and is currently pregnant. This is the first transgenic farm animal born in Greece.

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