Use of OPU-IVP and SO at CRV breeding program in the genomic era Erik Mullaart and Sybrand Merton

Impact on breeding program Holland Genetics Content Introduction Genomic Selection Impact on breeding program Selection procedures Delta Nucleus breeding program Application of OPU-IVP and SO Insire embryo program

CRV Worldwide

Genetic products of CRV 7 million units of dairy and beef semen CRV exports semen to over 50 countries Test program: 310 Holstein bulls 60 Red Holstein bulls 35 Fleckvieh bulls 35 Jersey bulls

Genomic selection

Which calf received the best genes from the parents?

Marker selection In early days: selection on individual genes (direct or via markers) Now: selection based on many markers spread across the whole genome, i.e. selection on all genetic material =GENOMIC SELECTION (or Genomewide Selection)

Genomic Selection, What does it bring us? Reliable breeding values at young age higher success rate to get top bulls lower costs for test proofs (60,000 markers for ~ € 200) shorter generation-interval Toelichting geven

Effects of Genomic Selection Increased genetic improvement (30-40%) Higher genetic level of test sires and embryo’s Wider portfolio of commercial bulls & higher level More outcross; less inbreeding Availability of packs of elite young sires (InSire)

Impact on breeding program; selection procedures

Use of GS in selection procedures; method implemented by CRV Step 1: Genotype proven bulls with reliable EBVs (reference sires) Find markers with an effect on EBVs Calculate marker effect for each trait Step 2: Genotype selection candidates Calculate GS Breeding Values (GBVs) with the marker effects Select the animals with the highest GBVs

3 full sisters on Delta. Pedigree ( Oman x Jocko Besne)

Impact on breeding program; Delta nucleus breeding program

16 years of ‘Delta nucleus’ Test heifers 4381 Selected donors 561 Before Genomic Selection large number of test heifers per year emphasis on independent test With Genomic Selection. limited number of test heifers optimal use of reproductive possibilities

After GS-selection 110 donor heifers Delta nucleus Selection of 800 calves GS selection After GS-selection 110 donor heifers MOET en OPU 25 - 30 embryos per heifer 100 selected heifers tested Selection of 10 -15 donor cows ET en OPU => 2500 embryos

Application of OPU-IVP and SO

Embryo production IVP sessions: 2000 / year SO: 300 / year 1.6 embryo/ session SO: 300 / year 4-5 embryos/ flush

Production Logistics 3 x SO at 13-17 months of age: 12 embryos Holland Genetics 3 x SO at 13-17 months of age: 12 embryos Inseminate at 17 months 17 x OPU at 18-20 months: 20 embryos Calving at 26 months ---------------- 32 embryos

Future Production Logistics Holland Genetics 2 x SO at 13-15 months of age: 8 embryos 10 x OPU at 16-17 months: 12 embryos Inseminate at 18 months 17 x OPU at 19-21 months: 20 embryos Calving at 27 months ---------------- 40 embryos

Working method: SO D -12 --- Start protocol FSH 1x D -11 --- FSH 2x D -10 --- FSH 2x + Prostaglandin with 5th FSH injection D - 9 --- FSH 2x + Prostaglandin with 6th FSH injection D - 8 --- FSH 1x D - 7 --- Heat + AI (2x 8-12 hr / 16-24 hr) D - 0 --- Flush + Prostaglandin + Progesterone application D +12 --- Remove PRID D +14 --- Heat D + 23 - D + 28 --- start SO again (D - 12)

Working method: OPU-IVP Day -1 Ovum Pick Up (OPU) In vitro maturation (IVM) Day 0 In vitro fertilization (IVF) Day 1 Start in vitro culture (IVC) Day 4 Cleavage evaluation Day 6 Prognoses embryo production Day 7 Embryo evaluation (fresh, frozen) Day 8 Final embryo evaluation (frozen)

Working method: OPU OPU twice a week (Mo, Thu) No superstimulation Holland Genetics Working method: OPU OPU twice a week (Mo, Thu) No superstimulation (para)veterinarian plus assistant Pregnant heifers and first parity donor cows

Working method: IVM Maturation for 16 - 28 h, 38.5°C, 20% O2 Holland Genetics Working method: IVM Maturation for 16 - 28 h, 38.5°C, 20% O2 Volume: transport and maturation in 0.5 ml in tubes

Working method: IVF Semen: Frozen-thawed Holland Genetics Working method: IVF Semen: Frozen-thawed Separation semen and extender by gradient centrifugation (90% / 45%) Volume: wells (0.5 ml), Incubation for 24 h, 38.5 °C, 20% O2

Working method: IVC Medium: SOFaaBSA Volume: wells (0.5 ml) Holland Genetics Working method: IVC Medium: SOFaaBSA Volume: wells (0.5 ml) Incubation for 6-8 days, 38.5°C, 5% O2

Working method: IVC Embryos Day 7 - 8, fresh / frozen Stage 4 - 9 Holland Genetics Working method: IVC Embryos Day 7 - 8, fresh / frozen Stage 4 - 9 (Morula - Hatched Blastocyst) Grade 1 and 2 IETS

Working method: Embryo transfer Holland Genetics Recipients Healthy, fertile recipients Natural cycling Day 6, 7, 8 or 9 of estrus cycle <5% rejected recipients 150 contracted farms

Working method: Logistics OPU / IVP Holland Genetics Working method: Logistics OPU / IVP OPU and SO location Transport COC’s Central IVP lab Transport of fresh embryos Recipient herds Farmers

Results: pregnancy rates Holland Genetics Results: pregnancy rates Fresh IVP embryos: 45% - 55% Frozen embryos: IVP 40% - 50% SO 50% - 60%

Results: Efficiency Oocytes: 8 9 Cleavage: 5,6 (70%) OPU-IVP MOET Oocytes: 8 9 Cleavage: 5,6 (70%) Embryo: 1,6 (20%) 5 (55%) Calf: 0,8 (10%) 3 (33%)

Insire embryo program

Number of InSire-embryos per year InSire-embryo donors Delta test heifers (IVP + SO) 2500 Delta donor cows (IVP + SO) 500 Total from Delta program = 3000 Marker selected Bullmothers in West-Europe. (e.g. France, Germany, Belgium, Denmark and the Netherlands 1000 Totaal 4000 InSire-embryos

Costs InSire-embryos InSire-embryos with option € 325,- InSire-embryos with segment option € 275,- CRV claims /buy 15-20% of the calves born male € 1000 female € 3000

Segmentation InSire-embryos Kg Milk % Protein Life span en health Top exterior

InSire-embryos offered by Internet 1) Random embryo selection (most actual combination) Choice of segment ( 4 ) Embryo price € 275 IVP fresh , IVP and SO frozen 2) Newest InSire embryos (most actual combination) Choice of combination Embryo price € 325 3) Available InSire embryos (in stock) IVP and SO frozen

2) Newest InSire embryos (most actual combination)

Benefits to the clients

Win-Win situation Client InSire embryos: - Top class calves for a relative low price CRV: - increased genetics of bulls and bull dams against an acceptable costs

Thank you for your attention!