Defined media optimization for in vitro culture of bovine somatic cell nuclear transfer (SCNT) embryos |
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| Affiliation: | 1. College of Veterinary Medicine, Northwest A & F University, Yangling, Shaanxi 712100, China;2. College of Life Science and Technology, Southwest University for Nationalities, Chengdu 610041, China;1. Division of Life Sciences, Korea Polar Research Institute, Incheon 21990, South Korea;2. Department of Biological Sciences, Inha University, Incheon 22212, South Korea;3. Institute of Life Science & Natural Resources, Korea University, Seoul 02841, South Korea;4. Department of Biology, Gangneung-Wonju National University, Gangneung 25457, South Korea;5. Consulting Engineering Office for Ecology, Radetzkystrasse 10, 5020 Salzburg, Austria;1. Department of Genetics and Developmental Biology, Institut Curie, Paris, France;2. CNRS UMR3215, Inserm U934, F-75248 Paris, France;1. Physiology & Climatology, Indian Veterinary Research Institute, Izatnagar, Bareilly, Uttar Pradesh 243122, India;2. Biochemistry Section, Indian Veterinary Research Institute, Izatnagar, Bareilly, Uttar Pradesh 243122, India;1. Department of Veterinary Biomedical Sciences, University of Saskatchewan, Saskatoon, Canada;2. Agriculture and Agri-Food Canada, Saskatoon Research and Development Center, Saskatoon, Canada;3. Department of Large Animal Clinical Sciences, University of Saskatchewan, Saskatoon, Canada;4. Reproductive Physiology, Toronto Zoo, Ontario, Canada;1. KEK Theory Center, High Energy Accelerator Research Organization (KEK), Japan;2. Graduate University for Advanced Studies (SOKENDAI), Tsukuba, Ibaraki 305-0801, Japan;3. Department of Physics, Tokyo Metropolitan University, Hachioji, Tokyo 192-0397, Japan |
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| Abstract: | The objective was to establish an efficient defined culture medium for bovine somatic cell nuclear transfer (SCNT) embryos. In this study, modified synthetic oviductal fluid (mSOF) without bovine serum albumin (BSA) was used as the basic culture medium (BCM), whereas the control medium was BCM with BSA. In Experiment 1, adding polyvinyl alcohol (PVA) to BCM supported development of SCNT embryos to blastocyst stage, but blastocyst formation rate and blastocyst cell number were both lower (P < 0.05) compared to the undefined group (6.1 vs. 32.6% and 67.3 ± 3.4 vs. 109.3 ± 4.5, respectively). In Experiment 2, myo-inositol, a combination of insulin, transferrin and selenium (ITS), and epidermal growth factor (EGF) were added separately to PVA-supplemented BCM. The blastocyst formation rate and blastocyst cell number of those three groups were dramatically improved compared with that of PVA-supplemented group in Experiment 1 (18.5, 23.0, 24.1 vs. 6.1% and 82.7 ± 2.0, 84.3 ± 4.2, 95.3 ± 3.8 vs. 67.3 ± 3.4, respectively, P < 0.05), but were still lower compared with that of undefined group (33.7% and 113.8 ± 3.4, P < 0.05). In Experiment 3, when a combination of myo-inositol, ITS and EGF were added to PVA-supplemented BCM, blastocyst formation rate and blastocyst cell number were similar to that of undefined group (30.4 vs. 31.1% and 109.3 ± 4.4 vs. 112.0 ± 3.6, P > 0.05). In Experiment 4, when blastocysts were cryopreserved and subsequently thawed, there were no significant differences between the optimized defined group (Experiment 3) and undefined group in survival rate and 24 and 48 h hatching blastocyst rates. Furthermore, there were no significant differences in expression levels of H19, HSP70 and BAX in blastocysts derived from optimized defined medium and undefined medium, although the relative expression abundance of IGF-2 was significantly decreased in the former. In conclusion, a defined culture medium containing PVA, myo-inositol, ITS, and EGF supported in vitro development of bovine SCNT embryos. |
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