1. Reproductive Biotech

2. Superovulation
treatment of the female with hormones so that more eggs are ovulated than normal

3. Superovulation alter normal level of FSH Follicle Stimulating Hormone
Typical results are ovulation of 10 or more eggs

4. Superovulation
recipient animals must be closely synchronized with the donor cow.
Must be at the same stage in their heat cycle

5. EmbryoTransfer
provides opportunity to introduce genetic material into populations of livestock while greatly reducing risk of transmission of diseases

6. ET Process super ovulate donor cow synchronize recipients
breed donor cow (A.I.) when in heat

7. ET Process
flush - remove fertilized eggs from donor before 7 days gestation
plant embryo in the recipient animal

8. Transfer to recip. Non surgical common
deposit embryo in uterus through cervix with AI straw and gun

9. non surgical
usually 70-80% success rate

10. Surgical incision in recip through flank
tip of uterine horn is exposed
cut open and embryo is deposited

11. Surgical
recip is sewn shut
less than 60-65% success rate.

12. ET
now allows for shipping of 500 registered holsteins in four sealed flasks in one seat on an airplane

13. Splitting Embryos remove embryo from cow 7 days after conception
collect unfertilized egg and cut into zona pellucida

14. Splitting Embryos
remove contents of unfertilized egg leaving the zona intact
similarly, cut into zona around embryo

15. Splitting Embryos
divide cells in embryo in half using a micro surgical blade
remove 1/2 of the cells and transfer them into empty zona

16. Splitting Embryos seal zona with microjolt
each 1/2 put into recipient animals

17. Cloning
6 days after conception, 32 cell embryo is removed from donor cow
recipient egg cell is taken from a slaughterhouse

18. Cloning recipient cells DNA is removed
one of the 32 cells is removed from the embryo

19. Cloning the single cell is inserted into recipient egg
recipient cell is given a microjolt - fuses grafted DNA to plasma membrane

20. Cloning
desired embryo will replace cells that are removed providing a continuing supply of DNA to be inserted into more recipient cells

21. Cloning
resulting offspring are cloned identical siblings

22. Sexing Embryos usually done by determining if embryo is a male
males have Y chromosome
H-Y antigen can be detected in male cells

23. Detection of Y
break open cells and stain chromosomes to identify presence of Y chromosome
Karyotyping
must examine at least 10 cells

24. Detection of Y 70% accurate use Y specific probe for ID
Y specific probe attaches to Y chromosome

25. Detection of Y can be detected by radioactivity or special light
85% accurate

26. H-Y Antigen make an antibody by injecting H-Y antigen into mice
attach dye onto the antibody for easy detection of male embryo

27. H-Y Antigen add antibody to embryo and if embryo is male it will glow
if it is female it will remain dark

28. Advantages does not kill cell no damage to embryo
fast - can be few hours or less than one hour old

29. Disadvantages
need high skilled technician
80% accurate

30. Freezing Embryos cooling embryos -5C or 40 F
can be stored several days
freezing -196 C or -320 F
must cool before freezing

31. Freezing Embryos
slowly cooled to -25C at a rate of .5C per minute and then plunged into liquid Nitrogen

32. Freezing
cryoprotectants are used to lower the freezing point so removal of water can be increased

33. Freezing Cryoprotectant used in cattle - glycerol
also help prevent cell membrane from becoming brittle
dilutes salt in cell

34. Thawing 500 C per minute complete is about 20 seconds
glycerol is removed and embryo is placed in successive solutions

35. Thawing
glycerol is removed and embryo is placed in successive solutions to re hydrate

36. Thawing
if done too slowly embryo will burst because large ice crystals will form
70-80% success rate

37. Animals Genetic Transformation two tools embryo micromanipultion
recombinant DNA technology

38. Genetic Trans.
Occurs when new genetic info is introduced into animals genome

39. Methods
ES - Embryonic Stem
EC - Embryonic Carcinoma cells

40. ES or EC
cells harboring the foreign gene are incorporated into blastocyst resulting in a mosaic animal

41. EC
potential for tumor formation

42. Retrovirus infection
retro. Carrying desired gene is used to transformhost
close to 100% efficient

43. Microinjection
of several hundred copies of DNA are injected into male pronucleus of one or two cell embryos

44. Microinjection inefficient mice - 10-15% are born
only 25% express new trait

45. Screening is it transgenic? Incorporation of gene expression of gene
germline incorporation

46. Agricultural Use
rabbit
sheep
pig
cow

47. The End !! :)