Royana: Successful Experience in Cloning the Sheep

Objective: This study describes our experiences in reproductive cloning using two different procedures resulting in birth of the first successfully cloned sheep in Iran and the Middle-East, nick-named "Royana". Materials and Methods: Abattoir-derived sheep oocytes were enucleated after in vitro maturation for 18-20hrs and then reconstructed by ear-derived sheep somatic cells using two different procedures of renucleation (subzonary, intracytoplasmic), embryo culture (coculture, sequential medium) and embryo transfer (intra fallopian, intra uterine). Pregnancy status and fetal development were followed regularly and elective cesarean was inducted on day 145 of pregnancy. Histopathological and genetical examinations were performed on either aborted and delivered clones for confirmation different aspects of cloning. Results: The two procedures were both efficient in producing early and/or advanced cloned embryos, establishing early and/or advanced stages of pregnancy till delivery. Four pregnancies were detected; one were failed at early pregnancy, one aborted on day 90, one was still born and the fourth delivered to a healthy male lamb nick named "Royana". Conclusion: Many different approaches have been developed for mammalian cloning which all are judged by their ultimate potency for establishment of successful pregnancies terminated to healthy/viable clones. As a preliminary study toward establishment of the technology, this study also successfully examined the competency of two procedures of somatic cell nuclear transfer (SCNT). However, the overall low efficiency of SCNT indicates that many different aspects of the technology remain to be dissolved.