34 Recent Advances in Cloning by Somatic Cell Nuclear Transfer in Camelids

Abstract

In addition to its application for production of elite males, racing champions, animals with the highest potential for milk production, and the prized beauty camels, somatic cell nuclear transfer (SCNT) can be utilised for the conservation of endangered wild Bactrian camels and vicunas by using the technique of interspecies SCNT (iSCNT). In the present study, embryos were reconstructed by using skin fibroblast cells from a Bactrian camel (Camelus bactrianus), a llama (Llama glama), and a dromedary camel (Camelus dromedarius) as donor karyoplasts and dromedary oocytes as recipient cytoplasts to evaluate in vitro and in vivo developmental potential of these embryos. Mature oocytes were collected from super-stimulated dromedary camels by ultrasound guided transvaginal ovum pick-up. Serum-starved skin fibroblast cells were injected into the perivitelline space of enucleated oocytes. The fibroblast cell and recipient cytoplasm were fused by 2 DC pulses of 100V for 15 µs each. Reconstructs were activated 1 to 1.5 h post-fusion with 5 µM ionomycin, followed by exposure to 6-DMAP for 4 h. The activated oocytes were then cultured at 38.5°C in an atmosphere of 5% CO₂, 5% O₂, and 90% N₂. The proportion of oocytes that cleaved was recorded on Day 3, and those that reached morula and blastocyst stages were recorded on Day 7 of culture. A lower blastocyst production rate was observed when llama fibroblast cells were used as karyoplast (13.3 ± 3.3) compared with those obtained from the embryos reconstructed with Bactrian (34.4 ± 3.9) and dromedary (32.2 ± 6.2) fibroblast cells; however, no difference was observed in their cell numbers. Out of 26 and 20 blastocysts from reconstructs with Bactrian and dromedary fibroblast cells, transferred to 23 and 12 synchronized dromedary recipients, 3 and 2 pregnancies were achieved at Day 60, respectively. One pregnancy from each group reached term and both (a Bactrian and a dromedary) calves were delivered normally after completing the gestation period. We demonstrated birth of a Bactrian calf conceived from the reconstructed embryo by iSCNT using the somatic cell from a Bactrian camel and enucleated dromedary oocyte. The present study also demonstrated that dromedary camel can be used as a surrogate to carry these pregnancies to term (Wani et al. 2017 PLoS One 12, e0177800; https://doi.org/10.1371/journal.pone.0177800). This study has opened doors for enhanced multiplication and preservation of the wild Bactrian camels, which are threatened with extinction, being the eighth most endangered large mammal on the planet.