title: Rex Rabbit Somatic Cell Nuclear Transfer with In Vitro-Matured Oocytes.
abstract: Somatic cell nuclear transfer (SCNT) requires large numbers of matured oocytes. In vitro-matured (IVM) oocytes have been used in SCNT in many animals. We investigated the use of IVM oocytes in Rex rabbit SCNT using Rex rabbit ovaries obtained from a local abattoir. The meiotic ability of oocytes isolated from follicles of different diameters was studied. Rex rabbit SCNT was optimized for denucleation, activation, and donor cell synchronization. Rex rabbit oocytes grew to the largest diameter (110 μm) when the follicle diameter was 1.0 mm. Oocytes isolated from <0.5-mm follicles lacked the ability to resume meiosis. More than 90% of these oocytes remained in the germinal vesicle (GV) stage after in vitro culture (IVC) for 18 h. Oocytes isolated from >0.7-mm follicles acquired maturation ability. More than 90% of these oocytes matured after IVC for 18 h. The developmental potential of oocytes isolated from >1-mm follicles was greater than that of oocytes isolated from 0.7- to 1.0-mm follicles. The highest activation rates for IVM Rex rabbit oocytes were seen after treatment with 2.5 μM ionomycin for 5 min followed by 2 mM 6-dimethylaminopurine (6-DMAP) and 5 μg/mL cycloheximide (CHX) for 1 h. Ionomycin induced the chromatin of IVM oocytes to protrude from the oocyte surface, promoting denucleation. Fetal fibroblast cells (FFCs) and cumulus cells (CCs) were more suitable for Rex rabbit SCNT than skin fibroblast cells (SFCs) (blastocyst rate was 35.6 ± 2.2% and 38.0 ± 6.0% vs. 19.7 ± 3.1%). The best fusion condition was a 2DC interval for 1 sec, 1.6 kV/cm voltages, and 40 μsec duration in 0.28 M mannitol. In conclusion, the in vitro maturation of Rex rabbit oocytes and SCNT procedures were studied systematically and optimized in this study.
authors.affiliation: 1 Key Laboratory of Embryo Development and Reproductive Regulation of Anhui Province, College of Biological and Food Engineering, Fuyang Teachers College , Fuyang City, Anhui Province 236037, China .
authors.forename: Yong
authors.initials: Y
authors.lastname: Liu
authors.affiliation: 2 Laboratory for Animal Reproduction and Embryology, College of Animal Science and Veterinary Medicine, Shandong Agricultural University , Tai'an City, Shandong Province 271018, China .
authors.forename: Huili
authors.initials: H
authors.lastname: Wang
authors.affiliation: 2 Laboratory for Animal Reproduction and Embryology, College of Animal Science and Veterinary Medicine, Shandong Agricultural University , Tai'an City, Shandong Province 271018, China .
authors.forename: Jinhua
authors.initials: J
authors.lastname: Lu
authors.affiliation: 2 Laboratory for Animal Reproduction and Embryology, College of Animal Science and Veterinary Medicine, Shandong Agricultural University , Tai'an City, Shandong Province 271018, China .
authors.forename: Yiliang
authors.initials: Y
authors.lastname: Miao
authors.affiliation: 2 Laboratory for Animal Reproduction and Embryology, College of Animal Science and Veterinary Medicine, Shandong Agricultural University , Tai'an City, Shandong Province 271018, China .
authors.forename: Xinyan
authors.initials: X
authors.lastname: Cao
authors.affiliation: 1 Key Laboratory of Embryo Development and Reproductive Regulation of Anhui Province, College of Biological and Food Engineering, Fuyang Teachers College , Fuyang City, Anhui Province 236037, China .
authors.forename: Ling
authors.initials: L
authors.lastname: Zhang
authors.affiliation: 1 Key Laboratory of Embryo Development and Reproductive Regulation of Anhui Province, College of Biological and Food Engineering, Fuyang Teachers College , Fuyang City, Anhui Province 236037, China .
authors.forename: Xiaoqing
authors.initials: X
authors.lastname: Wu
authors.affiliation: 1 Key Laboratory of Embryo Development and Reproductive Regulation of Anhui Province, College of Biological and Food Engineering, Fuyang Teachers College , Fuyang City, Anhui Province 236037, China .
authors.forename: Fengrui
authors.initials: F
authors.lastname: Wu
authors.affiliation: 1 Key Laboratory of Embryo Development and Reproductive Regulation of Anhui Province, College of Biological and Food Engineering, Fuyang Teachers College , Fuyang City, Anhui Province 236037, China .
authors.forename: Biao
authors.initials: B
authors.lastname: Ding
authors.affiliation: 1 Key Laboratory of Embryo Development and Reproductive Regulation of Anhui Province, College of Biological and Food Engineering, Fuyang Teachers College , Fuyang City, Anhui Province 236037, China .
authors.forename: Rong
authors.initials: R
authors.lastname: Wang
authors.affiliation: 2 Laboratory for Animal Reproduction and Embryology, College of Animal Science and Veterinary Medicine, Shandong Agricultural University , Tai'an City, Shandong Province 271018, China .
authors.forename: Mingjiu
authors.initials: M
authors.lastname: Luo
authors.affiliation: 1 Key Laboratory of Embryo Development and Reproductive Regulation of Anhui Province, College of Biological and Food Engineering, Fuyang Teachers College , Fuyang City, Anhui Province 236037, China .
authors.forename: Wenyong
authors.initials: W
authors.lastname: Li
authors.affiliation: 2 Laboratory for Animal Reproduction and Embryology, College of Animal Science and Veterinary Medicine, Shandong Agricultural University , Tai'an City, Shandong Province 271018, China .
authors.forename: Jinghe
authors.initials: J
authors.lastname: Tan
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001755:Blastocyst / Q000166:cytology; D054885:Cumulus Cells / Q000166:cytology* / Q000187:drug effects; D003513:Cycloheximide / Q000494:pharmacology; D005260:Female; D059471:In Vitro Oocyte Maturation Techniques; D015759:Ionomycin / Q000494:pharmacology; D008540:Meiosis; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000166:cytology* / Q000187:drug effects; D006080:Ovarian Follicle / Q000166:cytology; D010312:Parthenogenesis / Q000187:drug effects; D011817:Rabbits
chemical_list: D015759:Ionomycin; C001020:N(6),N(6)-dimethyladenine; D003513:Cycloheximide; D000225:Adenine