The cloning of equines by somatic cell nuclear trans- fer (SCNT) was first reported in 2003 (Galli et al., 2003; Woods et al., 2003), 6 years after the birth of Dolly the sheep (Wilmut et al., 1997). Amongst the mammalian spe- cies that have been cloned (Campbell et al., 2007), equines are among the last. In general, the application of assisted reproduction technologies in equines has lagged behind that of other livestock species (Galli et al., 2007) in the poor interest expressed by the industry (still today, for example, the thoroughbred does not allow artificial insemi- nation) and also the limited resources devoted to this kind of research and the few laboratories working in this area. In fact the first equine clone, a mule (Woods et al., 2003), was obtained from in vivo matured oocytes that were transferred back to the oviduct of a synchronized recipi- ent mare immediately after nuclear transfer and activation. This approach was successful because it limited to a mini- mum the time exposure to the in vitro environment. This approach of using in vivo matured oocytes was, however, not sustainable from a practical point of view. In contrast, the first cloned horses (Galli et al., 2003; Lagutina et al., 2005) and those that followed later (Hinrichs et al., 2006, 2007) were all derived from abattoir-recovered and in vitro matured oocytes that, following enucleation and embryo reconstruction, were cultured in vitro to the blastocyst stage prior to non-surgical embryo transfer to a recipient mare. This method could be applied only after the develop- ment of the procedures for in vitro maturation of oocytes, fertilization by ICSI, and in vitro culture of embryos capable of establishing normal pregnancies and generat- ing offspring (Galli et al., 2007; Hinrichs, 2010). In addi- tion to the refinement of the in vitro protocols for oocyte maturation and embryo culture, SCNT protocols had to be adapted and optimized for the equine, which, because of its anatomical and physiological features, provides a lim- ited number of oocytes per ovary when compared to cat- tle or pigs. The protocols currently applied for nuclear transfer are based on a zona-free system (Lagutina et al., 2007) to enhance cell fusion or a piezo-electric device (Westhusin et al., 2003) for enucleation and microinjec- tion of the nucleus directly into the oocyte. Reconstructed embryos also require a specific activation protocol (Hinrichs et al., 2006; Lazzari et al., 2002).
Cloning of Equines
GALLI, CESARE;LAZZARI, GIOVANNA
2014
Abstract
The cloning of equines by somatic cell nuclear trans- fer (SCNT) was first reported in 2003 (Galli et al., 2003; Woods et al., 2003), 6 years after the birth of Dolly the sheep (Wilmut et al., 1997). Amongst the mammalian spe- cies that have been cloned (Campbell et al., 2007), equines are among the last. In general, the application of assisted reproduction technologies in equines has lagged behind that of other livestock species (Galli et al., 2007) in the poor interest expressed by the industry (still today, for example, the thoroughbred does not allow artificial insemi- nation) and also the limited resources devoted to this kind of research and the few laboratories working in this area. In fact the first equine clone, a mule (Woods et al., 2003), was obtained from in vivo matured oocytes that were transferred back to the oviduct of a synchronized recipi- ent mare immediately after nuclear transfer and activation. This approach was successful because it limited to a mini- mum the time exposure to the in vitro environment. This approach of using in vivo matured oocytes was, however, not sustainable from a practical point of view. In contrast, the first cloned horses (Galli et al., 2003; Lagutina et al., 2005) and those that followed later (Hinrichs et al., 2006, 2007) were all derived from abattoir-recovered and in vitro matured oocytes that, following enucleation and embryo reconstruction, were cultured in vitro to the blastocyst stage prior to non-surgical embryo transfer to a recipient mare. This method could be applied only after the develop- ment of the procedures for in vitro maturation of oocytes, fertilization by ICSI, and in vitro culture of embryos capable of establishing normal pregnancies and generat- ing offspring (Galli et al., 2007; Hinrichs, 2010). In addi- tion to the refinement of the in vitro protocols for oocyte maturation and embryo culture, SCNT protocols had to be adapted and optimized for the equine, which, because of its anatomical and physiological features, provides a lim- ited number of oocytes per ovary when compared to cat- tle or pigs. The protocols currently applied for nuclear transfer are based on a zona-free system (Lagutina et al., 2007) to enhance cell fusion or a piezo-electric device (Westhusin et al., 2003) for enucleation and microinjec- tion of the nucleus directly into the oocyte. Reconstructed embryos also require a specific activation protocol (Hinrichs et al., 2006; Lazzari et al., 2002).I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
