Past, present and future of bovine in vitro embryo production in Denmark. Poul Hyttel and Lotte Strøbech University of Copenhagen

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1 Past, present and future of bovine in vitro embryo production in Denmark Poul Hyttel and Lotte Strøbech University of Copenhagen

2 Bovine in vitro embryo production in DK - looking back Royal Veterinary and Agricultural University (KVL) Departments of Reproduction and Anatomy First European in vitro produced calf 1987 Embryo Technology Center ( ) KVL, Danish Institute of Agricultural Sciences (DIAS) and DEMTEK A/S In vitro production to be developed for cattle breeding Obstructed by large offspring syndrome (LOS) and animal welfare considerations DIAS (>AU) and KVL (>KU) Fundamental aspects of oocyte maturation, fertilization and embryonic development

3 First European in vitro produced calf

4 First European in vitro produced calf 27 h 22 h 6 days 20 h

5 First European in vitro produced calf 27 h 22 h 6 days 20 h

6 Bovine in vitro embryo production in DK - looking back Royal Veterinary and Agricultural University (KVL) Departments of Reproduction and Anatomy First European in vitro produced calf 1987 Embryo Technology Center ( ) KVL, Danish Institute of Agricultural Sciences (DIAS) and DEMTEK A/S In vitro production to be developed for cattle breeding Obstructed by large offspring syndrome (LOS) and animal welfare considerations DIAS (>AU) and KVL (>KU) Fundamental aspects of oocyte maturation, fertilization and embryonic development

7 Large offspring syndrome 1997

8 Bovine in vitro embryo production in DK - looking back Royal Veterinary and Agricultural University (KVL) Departments of Reproduction and Anatomy First European in vitro produced calf 1987 Embryo Technology Center ( ) KVL, Danish Institute of Agricultural Sciences (DIAS) and DEMTEK A/S In vitro production to be developed for cattle breeding Obstructed by large offspring syndrome (LOS) and animal welfare considerations DIAS (>AU) and KVL (>KU) Fundamental aspects of oocyte maturation, fertilization and embryonic development

9 Gender-effect on embryonic development

10 Embryo culture 1999

11 Embryo culture 1999

12 Oocyte growth is related to competence Oocyte development

13 Electron microscopy of oocyte maturation Oocyte development

14 Electron microscopy of fertilization Fertilization

15 Bovine in vitro embryo production in DK - now AU, KU, EmbryoTrans Biotech GIFT: Genomic Improvement of Fertilization Traits in Danish and Brazilian Cattle (Innovation Fund Denmark) Key genes and genetic markers for most competent oocyte donors and embryo recipients AU, VikingGenetics, Trans Embryo Genetics Re-establishment of OPU and IVP for use in practice KU, University of Sheffield, Tartu University SEARMET Scientific Excellence in Animal Reproductive Medicine and Embryo Technology (EU) EU-based training in OPU and IVP

16 Granulosa transcriptome for finding the most competente oocyte dodors Cumulus cell RNA contributes to oocyte competence

17 Bovine in vitro embryo production in DK - now AU, KU, EmbryoTrans Biotech GIFT: Genomic Improvement of Fertilization Traits in Danish and Brazilian Cattle (Innovation Fund Denmark) Key genes and genetic markers for most competent oocyte donors and embryo recipients AU, VikingGenetics, Trans Embryo Genetics Re-establishment of OPU and IVP for use in practice KU, University of Sheffield, Tartu University SEARMET Scientific Excellence in Animal Reproductive Medicine and Embryo Technology (EU) EU-based training in OPU and IVP

18 Bovine in vitro embryo production in DK - future Biopsy for genomic selection 24 hours 24 hours 7 days 9 months Ovum pick-up from superior donor cows In vitro production of embryos Collection of embryonic biopsy for genomic selection according to current breeding measures Cryopreservation and transfer to optimal recipients Funding for future activities required!

19 The (in vitro fertilization) IVF laboratory at University of Copenhagen In vitro produced blastocysts 19

20 The scientific team behind the IVF laboratory at University of Copenhagen Birthe Avery, MD, PhD, DVSc Poul Hyttel, DVM, PhD, DVSc Lab Manager Master and PhD Students Birthe Avery, MD, PhD, DVSc Lotte Stroebech, DVM, PhD

21 Bovine in vitro production (IVP) of Embryos Two methods to retrieve oocytes: Ovum Pick Up OPU Or Collect ovaries from slaugtherhouse and aspirate oocytes

22 IVF Procedure in the Laboratory Aspiration (OPU) IVM (Day -1) IVF (Day 0) IVC (Day +1) Blastocyst score/ transfer or vitrification (Day 7-8) 22

23 Maturation (Day -1) Bovine Slaugter House ovaries Follicles above 2 mm and less than mm are aspirated In tubes containing 150 µl heparin and kept heated (30-32 C) 23

24 Fertilization IVF (Day 0) Good in vivo bull maybe not a good in vitro bull Mix 2 bulls + spermatocytes 24 h IVM 24 h IVF 2 PN 24

25 Culture IVC (Day 1) IVM (maturation) 24 h, 38.8 C, 6% CO 2 20% O 2 IVF (fertilization) 24 h, 38.8 C, 6% CO 2 20% O 2 IVC (embryo culture) 1 to 7-8 days 38.8 C, 6% CO 2 6% O 2 Low oxygen incubator 25

26 IVF Media from IVF Bioscience A product for every step of the in vitro production procedure

27 Thank you for your attention