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The objective of these experiments was to study bovine in vitro fertilization (IVF) conditions for blastocyst production using non-sexed sperm (Experiment 1) and sexed sperm (Experiment 2). For Experiment 1, in vitro-matured oocytes (N=707) were allocated to a 2 × 3 × 4 factorial design: time of co-incubation of gametes for fertilization (4 and 18 h), sperm dose (1, 0.33, and 0.11 × 10(6) frozen-thawed sperm/ml, and sperm source (four bulls). Pronuclear status was evaluated for a subset. Experiment 2 (N=2155 oocytes) was a 2 × 3 × 2 × 6 factorial design: sex of sperm (X and Y), sperm dose (1, 0.33, and 0.11 × 10(6) frozen-thawed sperm/ml), and sperm-sorting pressures (40 and 50 psi), replicated with sperm of six bulls. Presumptive zygotes were cultured 60 h in chemically defined medium-1 (CDM-1), and for 114 h in CDM-2. For Experiment 1, pronuclear formation, cleavage and blastocysts rates were greater for 1, and 0.33 × 10(6) than 0.11 × 10(6) sperm/ml (72 and 62 vs 42%; 89 and 81 vs 58%; and 21 and 17 vs 9%, respectively; all p<0.01); polyspermy was greater for 1, than 0.33 and 0.11 × 10(6) sperm concentrations (24 vs 2 and 0%; p<0.01). There were greater main effects (p<0.01) of pronuclear formation (69 vs 48%), polyspermy (13 vs 4%), and cleavage (63 vs 54%), at 18 than at 4 h of co-incubation of gametes (all p<0.01). For Experiment 2, cleavage and blastocyst rates were greater for 1 × 10(6) sperm/ml vs 0.33 and 0.11 (69%, 47%, and 30% cleavage and 30%, 14%, and 8% blastocysts) and 40 vs 50 psi (54% and 44% cleavage and 18% and 15% blastocysts) (p<0.01). A marked bull by fertilization sperm dose interaction was found for cleavage (p<0.05). The main conclusion was that the optimal sperm concentration for cleavage and producing blastocysts via IVF with sexed sperm was considerably higher and more variable among bulls than for unsexed sperm.

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