A Transgenic animal is one that carries a foreign gene that has been deliberately inserted into its genome. The foreign gene are inserted into the germ line of the animal, so it can be transmitted to the progeny.

1. Role Of Transgenic Animal In
Target Validation
Nikita Bankoti
M.Pharm 2nd Sem
Pharmacology

2. Definition
•A Transgenic animal is one that carries a foreign gene that has been deliberately inserted into its
genome.
•The foreign gene are inserted into the germ line of the animal, so it can be transmitted to the
progeny.
•Transgenic animals are animals that are genetically altered to have traits that mimic symptoms of
specific human pathologies.
•They provide genetic model of various human disease which are important in understanding
disease and development of new target.

3. Terminologies
•Transgenesis: It is the process of introducing an exogenous gene- called a transgene-
into a living organism so that the organism will exhibit a new property and transmit that
property to its offspring.
•Transgene: It is the name given to the introduced gene or genetic material that has been
transferred naturally or by any of a number of genetic engineering techniques from one
organism to another.
•Knock in: It is a genetic engineering method where a specific gene or DNA sequence is
inserted or “knocked in” at a targeted location in the genome of an organism.
•Knock out: It is a genetic engineering technique used to disable or “knock out” a
specific gene in an organism’s genome. This is typically achieved by introducing a
mutation or deleting a section of the gene, rendering it non-functional.

4. Advantages
•Increased growth rate
•Improved disease resistance
•Increased muscle mass
•Improved nutritional quality
•Used as disease model
•Xenotransplantation

5. Disadvantage
•Inserted gene has multiple functions
•Breeding problems
•Sometimes lead to mutagenesis and functional disorders
•Low survival rate of transgenic animals
•Very expensive

6. Production Of Transgenic Animal
Step 1- construction of a transgene
Transgene is made up of 3 parts:
•Promoter
•Gene to be expressed
•Termination sequence

7. Step 2- introduction of foreign gene into the animal
•DNA Microinjection method
•Embryonic stem cell-mediated gene transfer
•Retrovirus mediated gene transfer
Step 3- screening for transgenic positives
•Transgenic progenies are screened by PCR to examine the site of incorporation of the gene.
•Some transgenes may not be expressed if integrated into a transcriptionally inactive site.
Step 4- further animal breeding is done to obtain maximal expression
•Heterozygous offsprings are mated to form homozygous strains.

8. 1.DNA Microinjection Method
•This procedure involves delivery of foreign gene to the living cell by using fine glass micro
pipettes and micro injection.
•A female animal is supur-ovulated and eggs are collected.
•The eggs are fertilized by in vitro method.
•The transgene containing solution is injected into the male pronucleus using micropipette.
•Eggs with the transgenes are kept overnight in an incubator to develop to a 2 cell stage.
•The eggs are then implanted into the uterus of a pseudo pregnant female(female which has been
mated with a vasectomized male the previous night).
•Live birth of transgenic animal
•The first successful DNA microinjection was carried in mouse.

9. DNA Microinjection Method
b. Transgene microinjection
a. Eggs collection and fertilization
c. Transgenic zygote
transfer into host
d. Implanted female
pregnancy and delivery

10. 2.Embryonic Stem Cell Mediated Gene
Transfer
•Transgenic animals can be created by manipulating embryonic stem cells.
• ESC are obtained from the inner cell mass of the blastocyst.
•Transgene is incorporated into ESC by-
-Microinjection
-by retro virus
-By electroporation
•Transgenic stem cells are grown in vitro.
•Then they are inserted into a blastocyst and implanted into a host’s uterus to grow
normally.(foster mother)
•Harvest the inner cell mass (ICM) from 3 day old blastocysts.

12. 3.Retrovirus-Mediated Gene Transfer
A retrovirus is a virus that carries its genetic material in the form of RNA rather than DNA.
This method involves-
•Retrovirus used as vectors to transfer genetic material into host cell, resulting in a chimera( an
organism or tissue that contains at least two different sets of DNA, most often originating from
the fusion of as many different zygotes), an organism consisting of tissues or parts of diverse
genetic constitution.
•Chimeras are inbred (breeding between closely related individuals) for as many as 20 generations
until homozygous (carrying the desired transgene in every cell) transgenic offspring are born.

13. Retrovirus-mediated gene transfer

14. Role of transgenic animals in target validation
1. Target identification
•Suppose we identify disease i.e., Alzheimer
•Translated this disease model into animal
•Animal model of Alzheimer’s disease ( streptozotocin used to induce beta amyloid protein in
hippocampus)
•Target identification by proteomics and genomics
•Overexpression of APP protein is responsible for disease.

15. 2. Target validation
•Validate that really APP (amyloid precursor protein) is responsible for Alzheimer’s disease.
•Deliver mutant amyloid precursor protein gene into animal.
•Progenies having mutant APP gene overexpressed ( resulted in Alzheimer’s disease)
•Target validated

16. 3. Identifying novel genes that are potential drug target.
•Target interaction- interaction with a perfect target should result in the prevention or regression of
disease.
•Not lead in serious target related side effects.
•Target should belong to a class of molecules that could be affected by small orally available
molecule.
4. Tissue-specific targeting
•Transgenic animals can express or supress genes selectively in specific tissue or cell types
relevant to the disease under investigation.
•This allow researchers to assess the role of the target gene in specific tissues and organs, helping
to elucidate its contribution to disease pathology.

17. 5. Phenotypic Characterization
•Transgenic animals can be phenotypically characterized to evaluate disease- relevant phenotypes such
as change in behaviour, physiology, or biochemical markers. By correlating phenotypic changes with
the expression or manipulation of the target gene, researchers can validate its relevance to the disease
phenotype.
6. Functional Genomics
•Transgenic animals contribute to functional genomics studies, where large-scale gene manipulation
helps uncover novel targets.
7. Disease modeling
•Transgenic animals can be used as valuable model for studying human disease.
•By introducing disease-associated genetic mutations into animals or altering the expression of specific
genes, researchers can mimic aspects of human diseases in animals and investigate disease
mechanisms and investigate disease mechanism and potential therapeutic interventions.

18. Examples-

20. Examples of transgenic animal

23. “THANK YOU”