Biotechnology: Status and UsesReproductive Biotechnology'sIn Vitro Production of EmbryosIn Vivo Embryo ProductionStimulating Follicular DevelopmentNon-Surgical Embryo FlushStage of Embryo at RecoveryET - SchemeEmbryo SexingHy-AntigenPowerPoint PresentationPCR Approach to Embryo SexingSperm Gender SelectionPERCENT DNA DIFFERENCES BETWEEN X and Y CHROMOSOMESlide 15Slide 16Fetal SexingCloningCloning by Nuclear TransferNuclear Cloning to Produce Stem CellsStem Cells Grown in Culture and Allowed to DifferentiateBenefits of Stem Cell ResearchGenetic ManipulationTransgenic for Growth HormoneGene Transfer Using Micro-Injection of PronucleiGene Transfer Using Viral TransfectionBiotechnology: Status and UsesBiotechnology: Status and UsesAnimal Science 434John J. ParrishAnimal Science 434John J. ParrishReproductive Biotechnology'sReproductive Biotechnology's•Artificial insemination•In vitro embryo production•In vivo embryo production•Embryo transfer•Gender selection•Stem Cells•Genetic engineering•Cloning•Artificial insemination•In vitro embryo production•In vivo embryo production•Embryo transfer•Gender selection•Stem Cells•Genetic engineering•CloningIn Vitro Production of EmbryosIn Vitro Production of Embryos•Oocyte isolation and maturation•Sperm preparation•Sperm capacitation•Fertilization•Embryo Development•Oocyte isolation and maturation•Sperm preparation•Sperm capacitation•Fertilization•Embryo DevelopmentIn Vivo Embryo ProductionIn Vivo Embryo Production•Normal cycling female–horses•Superovulated female–Cattle–Sheep–Goats–Deer–Humans•Normal cycling female–horses•Superovulated female–Cattle–Sheep–Goats–Deer–HumansStimulating Follicular DevelopmentStimulating Follicular DevelopmentEstrusEstruseCG orFSHeCG orFSHProgesterone From C.L.Progesterone From C.L.First FollicularWaveFirst FollicularWave10-1210-12MultipleOvulationsMultipleOvulationsPGF2PGF2Non-Surgical Embryo FlushNon-Surgical Embryo FlushDay 6 or 7Day 6 or 7Stage of Embryo at Recovery Stage of Embryo at Recovery Tight Morula (day 5 - 7)Tight Morula (day 5 - 7)Early Blastocyst (day 7 - 8)Early Blastocyst (day 7 - 8)Blastocyst (day 7 - 9)Blastocyst (day 7 - 9)ET - SchemeET - SchemeEmbryo SexingEmbryo Sexing•Hy Antigen–Associated with male cells•PCR and Detection of Y and X DNA•Hy Antigen–Associated with male cells•PCR and Detection of Y and X DNAHy-AntigenOctopus Springs - Yellowstone National ParkOctopus Springs - Yellowstone National ParkHome of Thermus aquaticus - Taq PolymeraseHome of Thermus aquaticus - Taq PolymerasePCR Approach to Embryo SexingXYF M F F F F F FSperm Gender SelectionSperm Gender SelectionSelection of X or Y spermSelection of X or Y spermPERCENT DNA DIFFERENCES BETWEEN X and Y CHROMOSOMEPERCENT DNA DIFFERENCES BETWEEN X and Y CHROMOSOMEHuman 2.9Cattle 3.8Chinchilla 7.5Turkey 0Human 2.9Cattle 3.8Chinchilla 7.5Turkey 0X Chromosome has more DNA!!!X Chromosome has more DNA!!!++--+-LLAASSEERR90°0°XX ssoorrttYY ssoorrttSpermStainedWith DNASensitiveFluorescentDyeSpermStainedWith DNASensitiveFluorescentDyeFlow Cytometer Separation of X and Y SpermFlow Cytometer Separation of X and Y SpermSperm Gender SelectionSperm Gender Selection•Flow Cytometry–Only method that works!–Very few sperm recovered–Reduced fertility–Expense will limit use•Flow Cytometry–Only method that works!–Very few sperm recovered–Reduced fertility–Expense will limit useFetal SexingORUltrasound EvaluationUltrasound Evaluation•Day 55 - 65•Day 55 - 65CloningCloningSplit morulaSplit morulaCloning by Nuclear TransferCloning by Nuclear TransferCycles are limitedOnly 3 - 4 cyclesNuclear Cloning to Produce Stem CellsNuclear Cloning to Produce Stem CellsCells of InterestGrown in CultureDish Cells of InterestGrown in CultureDish Mature Oocyte enucleated and cell of interest fusedMature Oocyte enucleated and cell of interest fusedEmbyro??DevelopsEmbyro??DevelopsInner Cell Mass Cells isolated and grown in cultureInner Cell Mass Cells isolated and grown in cultureDifferentiateDifferentiatePotential Tissue:•Neural•Adipose•Connective•HeartPotential Tissue:•Neural•Adipose•Connective•HeartStem Cells Grown in Culture and Allowed to DifferentiateStem Cells Grown in Culture and Allowed to DifferentiateQuickTime™ and aSorenson Video 3 decompressorare needed to see this picture.http://www.ansci.wisc.edu/jjp1/as434/powerpoint/fa07/stem_cells.movBenefits of Stem Cell ResearchBenefits of Stem Cell Research1. Regenerative medicine and tissue engineeringIn vitro systems: drug discovery, toxicology, diagnostic assays and cell culture reagentsBiomedical research: unravel mechanisms of disease and human development1. Regenerative medicine and tissue engineeringIn vitro systems: drug discovery, toxicology, diagnostic assays and cell culture reagentsBiomedical research: unravel mechanisms of disease and human developmentGenetic ManipulationGenetic ManipulationTransgenic for Growth HormoneTransgenic for Growth HormoneGene Transfer Using Micro-Injection of PronucleiGene Transfer Using Micro-Injection of PronucleiLess than 1% efficiencyGene Transfer Using Viral TransfectionGene Transfer Using Viral TransfectionBetter success but left with potential for viral replicationBetter success but left with potential for viral
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