In vitro oocyte culture and somatic cell nuclear transfer used to produce a live-born cloned goat.
The use of an in vitro culture system was examined for production of somatic cells suitable for nuclear transfer in the goat. Goat cumulus-oocyte complexes were incubated in tissue culture medium TCM-199 supplemented with 10% fetal bovine serum (FBS) for 20 h. In vitro matured (IVM) oocytes were enucleated and used as karyoplast recipients. Donor cells obtained from the anterior pituitary of an adult male were introduced into the perivitelline space of enucleated IVM oocytes and fused by an electrical pulse. Reconstituted oocytes were cultured in chemically defined medium for 9 days. Two hundred and twenty-eight oocytes (70%) were fused with donor cells. After in vitro culture, seven somatic cell nuclear transfer (SCNT) oocytes (3%) developed to the blastocyst stage. SCNT embryos were transferred to the oviducts of recipient females (four 8-cell embryos per female) or uterine horn (two blastocysts per female). One male clone (NT1) was produced at day 153 from an SCNT blastocyst and died 16 days after birth. This study demonstrates that nuclear transferred goat oocytes produced using an in vitro culture system could develop to term and that donor anterior pituitary cells have the developmental potential to produce term offspring. In this study, it suggested that the artificial control of endocrine system in domestic animal might become possible by the genetic modification to anterior pituitary cells.
Ohkoshi
Katsuhiro
K
Developmental Biology Department, National Institute of Agrobiological Sciences, Ibaraki, Japan.
Takahashi
Seiya
S
Koyama
Shin-Ichiro
S
Akagi
Satoshi
S
Adachi
Noritaka
N
Furusawa
Tadashi
T
Fujimoto
Jun-Ichiro
J
Takeda
Kumiko
K
Kubo
Masanori
M
Izaike
Yoshiaki
Y
Tokunaga
Tomoyuki
T
D000015:Abnormalities, Multiple / Q000662:veterinary; D000818:Animals; D001831:Body Temperature; D001835:Body Weight; D002478:Cells, Cultured; D019976:Cloning, Organism / Q000009:adverse effects / Q000379:methods*; D003470:Culture Media; D004272:DNA, Mitochondrial; D004624:Embryo Transfer; D005260:Female; D005838:Genotype; D006041:Goats; D008285:Major Histocompatibility Complex; D008297:Male; D053652:Nuclear Transfer Techniques; D009865:Oocytes; D010053:Ovary / Q000166:cytology; D010903:Pituitary Gland, Anterior / Q000166:cytology*
D003470:Culture Media; D004272:DNA, Mitochondrial