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Improvement of canine somatic cell nuclear transfer procedure

Cited 23 time in Web of Science Cited 25 time in Scopus
Authors
Jang, Goo; Oh, H.J.; Kim, M.K.; Fibrianto, Y.H.; Hossein, M.S.; Kim, H.J.; Kim, J.J.; Hong, S.G.; Park, J.E.; Kang, S.K.; Lee, Byeong Chun
Issue Date
2007-10-18
Publisher
Elsevier
Citation
Theriogenology 2008;69:146-54
Keywords
Canine embryoSCNTParthenogenetic activationEmbryo transferIntergeneric
Abstract
The purpose of the present study on canine somatic cell nuclear transfer (SCNT) was to evaluate the effects of fusion strength, type of activation, culture media and site of transfer on developmental potential of SCNT embryos. We also examined the potential of enucleated bovine oocytes to serve as cytoplast recipients of canine somatic cells. Firstly, we evaluated the morphological characteristics of in vivo-matured canine oocytes collected by retrograde flushing of the oviducts 72 h after ovulation. Secondly, the effectiveness of three electrical strengths (1.8, 2.3 and 3.3 kV/cm), used twice for 20 μs, on fusion of canine cytoplasts with somatic cells were compared. Then, we compared: (1) chemical versus electrical activation (a) after parthenogenetic activation or (b) after reconstruction of canine oocytes with somatic cells; (2) culture of resulting intergeneric (IG) embryos in either (a) mSOF or (b) TCM-199. The exposure time to 6-DMAP was standardized by using bovine oocytes reconstructed with canine somatic cells. Bovine oocytes were used for SCNT after a 22 h in vitro maturation interval. The fusion rate was significantly higher in the 3.3 kV/cm group than in the 1.8 and 2.3 kV/cm treatment groups. After parthenogenesis or SCNT with chemical activation, 3.4 and 5.8%, respectively, of the embryos developed to the morula stage, as compared to none of the embryos produced using electrical activation. Later developmental stages (8–16 cells) were transferred to the uterine horn of eight recipients, but no pregnancy was detected. However, IG cloned embryos (bovine cytoplast/canine somatic cell) were capable of in vitro blastocyst development. In vitro developmental competence of IG cloned embryos was improved after exposure to 6-DMAP for 4 h as compared to 0, 2 or 6 h exposure, although the increase was not significantly different among culture media. In summary, for production of canine SCNT embryos, we recommend fusion at 3.3 kV/cm, chemical activation, culture in mSOF medium and transfer of presumptive zygotes to the oviduct of recipient animals. The feasibility of IG production of cloned canine embryos using bovine cytoplasts as recipient of canine somatic cells was demonstrated.
ISSN
0093-691X
Language
English
URI
http://hdl.handle.net/10371/7620
DOI
https://doi.org/10.1016/j.theriogenology.2007.08.022
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College of Veterinary Medicine (수의과대학)Dept. of Veterinary Medicine (수의학과)Journal Papers (저널논문_수의학과)
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