Somatic cell nuclear transfer (SCNT) is an alternative potential tool for the conservation of endangered. In this study, somatic cells were collected from a purebred 9-month-old male mastiff and an 11-month-old female mastiff. Oocytes that had been matured in vivo were retrieved from outbred dogs by laparoscopy. We used cycling cells as donor cells for SCNT. A total of 289 oocytes were reconstructed with each male or female somatic cell and then fused/activated simultaneously by electrical stimulation. Finally, 224 embryos were transferred to 16 recipients that had been synchronized naturally. The efficiency of delivery of cloned dogs (7.1%) was threefold higher than in previous reports. Moreover, one surrogate delivered four identical cloned female Tibetan Mastiff puppies; another three surrogates each delivered triplets. Microsatellite analysis demonstrated the genotypic identity of the cloned puppies. Thus, our study has demonstrated techniques that improve significantly the overall efficiency of SCNT in the canine species.

Key words: Cloned mastiffs, Cycling adult skin fibroblasts, Plk gene, SCNT