32 CLONING OF ELITE QUARANTINE SNIFFING DOG BY SOMATIC CELL NUCLEAR TRANSFER

Abstract

Somatic cell nuclear transfer (SCNT) in assisted reproductive technologies has been considered for the conservation of valuable or endangered animals. Dogs that were originally bred for hunting, such as beagles, have an exceptional ability to detect a particular smell from many others. For that reason, the beagles have been used to detect quarantine risk items from a wide range of goods in assorted luggage without scaring or disrupting the passengers. Though very useful and highly in need, elite quarantine sniffing beagles with excellent abilities are rare; much time, effort, and money are required in producing them. Here, we have applied SCNT for propagation of elite quarantine sniffing dogs to save time and economic burden. Ear fibroblasts from a 10-year-old adult male elite quarantine sniffing beagle were isolated and cultured in vitro as donor cells. For SCNT, in vivo-matured oocytes, obtained by flushing the uterine tubes of oocyte donors (mixed breed), were used. The oocytes were enucleated, microinjected with donor cells, fused by electrical stimulation, and activated chemically. Reconstructed oocytes were surgically transferred into the uterine tube of naturally synchronous recipient females. A total of 212 activated cloned embryos were transferred into 12 female recipient dogs and 4 recipients became pregnant. The 4 pregnant recipients delivered 4 pups through caesarean section or natural delivery, but 1 died right after birth and did not show an abnormality. Other live puppies exhibited normal phenotypes; their appearance was similar to that of the donor dog. All cloned pups were genetically identical to the donor dog and their mitochondrial DNA was from their oocyte donor dogs. When the cloned pups were 16 weeks old, we conducted a Volhard test, which is commonly used to describe the following puppy aptitudes: social attraction, following, restraint, social dominance, elevation dominance, retrieving, touch sensitivity, sound sensitivity, and sight sensitivity. Dog behavior data on differences in transcript abundance were analyzed by a general linear mixed model. The 3 cloned pups showed similar behavioral tendencies. The present study demonstrates that NT technique using donor cell derived from 1 elite quarantine sniffing dog is useful to produce a large number of quarantine sniffing dogs.

This study was supported by RDA (no. PJ0089752012), RNL Bio (no. 550-20120006), IPET (no. 311062-04-1-SB010), Research Institute for Veterinary Science, Nestlé Purina Korea, and TS Corporation.