1. Rainbow
and her
clone, CC,
from TAMU
TEKS Ag: 130.2.12.D
130.3.6.A-D
130.7.6.B
130.7.12.A

2.  What is cloning?
 DNA Cloning
 Transferring DNA into a host cell for recombinant
DNA molecule
 Reproductive Cloning
 Generating a new animal with identical nuclear DNA
 Therapeutic Cloning aka “Embryo Cloning”
 Producing human embryos for research

4.  Form of Asexual Reproduction

6.  Semen Collection
 Assess potential fertility of male
 Short or long term storage before AI
 Ovum Collection
 Ovarian hyperstimulation – pharmacologically
stimulated to mature follicles
 Embryo Collection
 Superovulation to release multiple oocytes that can
be inseminated

7.  Performed using artificial vagina, dummy
mounting or electroejaculation
 Used fresh on the same day, stored for up to 2
days at refrigerator temperature, or frozen for
long periods of time
 Frozen semen mixed with diluter and extender
 Determine concentration
and motility
 Used for AI or IVF

8.  Intracervical
 Intrauterine
 Intrauterine tuboperitoneal
 Intratubal

9. PROS CONS
 Inseminate more females  Human Error
 Avoid injury or physical  Some equine breeding
breeding problems
prohibits AI
 Control paternity
 Requires specific facilities
 Use semen from males
separated from females by and skilled labor
time or distance  Increases costs of
 Decrease costs of keeping breeding
male
 Increase revenue of
offspring
 Increase reproductive
success
 Decrease contamination
and disease transmission

10.  Less than 10% of beef cattle; more than 90% of
dairy cattle
 More than 80% of the swine industry
 0% of the chicken industry
 100% of the turkey industry
 Selectively bred turkey increased the breast size so
male turkeys (Toms) are physically incapable of
mating

11.  Superovulation by injection of Equine
Chorionic Gonadotropin and prostaglandin F2
alpha or by injection of Follicle Stimulating
Hormone
 Egg can be used for in vitro fertilization
 Egg can be used for cloning
 Genetic material is removed to make an enucleated
cell

12.  Creating a clone embryo with a donor nucleus
 Somatic Cell – any cell in the organism that is not a
gamete
 Somatic cell from adult organism is collected
and the nucleus is transferred into the host
enucleated egg
 Host cell reprograms donor
nucleus and divides
mitotically to blastocyst
( about 5 days, about 100
cells)

13. 1. Oocytes collected in stage of Meiosis II
2. Chromosomes of oocytes removed by
micromanipulation to create cytoplast
3. Diploid 2N somatic cells from donor tissue
sample in G0 or G1 of Mitosis
4. Somatic cell is combined with cytoplast by
direct injection or by fusion using electric
pulse
5. Recombined ooctye artificially activated
6. Ooctye begins embryonic development and is
transferred to surrogate

15.  Multiple ovum after hormonal ovary stimulation are
inseminated, typically by AI
 Embryos are non-surgically collected by flushing with
a collection line and caught in a filter
 Non-surgical Embryo Transfer: embryos packaged into
straws and implanted in surrogate dam midway up the
uterine horn
 Surgical Embryo transfer: embryos are transferred with
laparoscopy by penetration of abdominal cavity
 Embryo transfer:
Fresh: approx. 65% conception rate
Frozen: approx. 40% conception rate

16.  Holstein Donor cow with calves carried by beef cattle surrogate dams

17.  After fertilization, at the 6 to 8 cell stage, the
embryo is split into two
 These are used for embryo transfer
 Both embryos then continue to develop into
two identical twins

18. PRO CON
 Acquiring better genetic  Decreases genetic
traits in offspring diversity
 Producing multiple  Increased costs of
high quality offspring breeding
 Increasing desired traits  Unresolved ethical
in a population issues
 Offspring traits are pre-  Increased chance of
determined birth defects
 Preserve endangered or  Does not ensure exact
extinct species replica

19. Idaho Gem, 1st cloned equine, 2003
Dolly, 1st cloned mammal, 1996
Millie, 1st cloned CC, 1st cloned
Snuppy, 1st cloned cat, 2001
cow, 2000
dog, 2005

20.  Semen extended with glycerol can
be frozen with liquid nitrogen (320
degrees below 0)
 Frozen semen is stored in straws
which can be thawed to directly inseminate a
female
 Embryo cryopreservation stores pre-implantation
stage embryos using slow cooling technique
 Frozen embryos in ethylene glycol can be thawed
for direct transfer
 Frozen embryos in 10% glycerol must be thawed
and rinsed multiple times before transfer

21.  Cryogenics is the engineering of tools to keep
very low temperatures
 Cryonics is the use of cryogenics in medicnal
cryopreservation
 Frozen semen and embryos for AI and embryo
transfer

22.  Synchronization breeding – using hormones to
sync up females coming into heat; typically
used for AI or embryo transfer
 Superovulation to promote twinning
 Automatic milking systems – electronic
identification of dairy cattle allows sorting of
cows for milking and breeding

23.  Selecting semen:
beef industry
prefers male
calves while
dairy industry
prefers female
calves

24.  Intravaginal progesterone insert to synchronize
estrus
 Helps to improve results of AI and estrus
synchronization
 Used in combination with gonadotropin
releasing hormone and prostaglandin F2 alpha
 Removes need to detect heat
 Increases pregnancy rate