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Biotechnology: Status and Uses. Animal Science 434 John J. Parrish. Reproductive Biotechnology's. Artificial insemination In vitro embryo production In vivo embryo production Embryo transfer Gender selection Stem Cells Genetic engineering Cloning. In Vitro Production of Embryos.

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Biotechnology: Status and Uses

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Biotechnology status and uses l.jpg

Biotechnology: Status and Uses

Animal Science 434

John J. Parrish


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Reproductive Biotechnology's

  • Artificial insemination

  • In vitro embryo production

  • In vivo embryo production

  • Embryo transfer

  • Gender selection

  • Stem Cells

  • Genetic engineering

  • Cloning


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In Vitro Production of Embryos

  • Oocyte isolation and maturation

  • Sperm preparation

  • Sperm capacitation

  • Fertilization

  • Embryo Development


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In Vivo Embryo Production

  • Normal cycling female

    • horses

  • Superovulated female

    • Cattle

    • Sheep

    • Goats

    • Deer

    • Humans


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PGF2a

eCG orFSH

Multiple

Ovulations

Estrus

Estrus

Progesterone

From C.L.

First Follicular

Wave

10-12

Stimulating Follicular Development


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Non-Surgical Embryo Flush

Day 6 or 7


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Stage of Embryo at Recovery

Tight Morula (day 5 - 7)

Early Blastocyst (day 7 - 8)

Blastocyst (day 7 - 9)


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ET - Scheme


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Embryo Sexing

  • Hy Antigen

    • Associated with male cells

  • PCR and Detection of Y and X DNA


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Hy-Antigen


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Octopus Springs - Yellowstone National Park

Home of Thermus aquaticus - Taq Polymerase


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PCR Approach to Embryo Sexing

Y

X

F M F F F F F F


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Sperm Gender Selection

Selection of X or Y sperm


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PERCENT DNA DIFFERENCES BETWEEN X and Y CHROMOSOME

Human 2.9

Cattle 3.8

Chinchilla 7.5

Turkey 0

X Chromosome has more DNA!!!


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Flow Cytometer Separation of X and Y Sperm

Sperm

Stained

With DNA

Sensitive

Fluorescent

Dye

9

L

A

S

E

R

-

+

+

-

Y

s

o

r

t

X

s

o

r

t


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Sperm Gender Selection

  • Flow Cytometry

    • Only method that works!

    • Very few sperm recovered

    • Reduced fertility

    • Expense will limit use


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Fetal Sexing

OR

Ultrasound Evaluation

  • Day 55 - 65


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Cloning

Split morula


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Cloning by Nuclear Transfer

Cycles are limited

Only 3 - 4 cycles


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Nuclear Cloning to Produce Stem Cells

Cells of InterestGrown in Culture

Dish

Mature Oocyte enucleated and cell of interest fused

Embyro??

Develops

  • Potential Tissue:

  • Neural

  • Adipose

  • Connective

  • Heart

Differentiate

Inner Cell Mass Cells isolated and grown in culture


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Stem Cells Grown in Culture and Allowed to Differentiate

http://www.ansci.wisc.edu/jjp1/as434/powerpoint/fa07/stem_cells.mov


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Benefits of Stem Cell Research

  • Regenerative medicine and tissue engineeringIn vitro systems: drug discovery, toxicology, diagnostic assays and cell culture reagentsBiomedical research: unravel mechanisms of disease and human development


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Genetic Manipulation


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Transgenic for Growth Hormone


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Gene Transfer Using Micro-Injection of Pronuclei

Less than 1% efficiency


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Gene Transfer Using Viral Transfection

Better success but left with potential for viral replication


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