Here are professor-level, university exam notes in
proper notes format for both units. These are
structured with definitions, headings, sub-points,
mechanisms, examples, and applications so you can
directly write in exams.
UNIT 1: ANIMAL CELL CULTURE AND
APPLICATIONS
1. Definition of Animal Cell Culture
Animal cell culture is the in vitro growth and
maintenance of animal cells, tissues, or organs in
artificial controlled conditions, outside the living
organism.
Key requirements
● Sterile environment
● Nutrient culture medium
● Controlled temperature (37°C for mammalian
cells)
● Proper pH (7.2–7.4)
● CO₂ incubator (5% CO₂)
● Growth factors and hormones
Importance
● Study of cell biology
● Vaccine production
● Protein production
● Cancer research
● Drug testing
2. Primary Cells and Cell Lines
Primary Cells
Definition:
Cells directly isolated from animal tissues and
cultured for the first time.
Sources
● Liver
● Kidney
● Skin
● Muscle
Characteristics
● Closely resemble natural cells
● Limited lifespan
● Anchorage dependent
● Contact inhibition present
● Genetically normal
Advantages
● Physiologically accurate
● Useful in toxicity testing
Disadvantages
● Limited growth
● Difficult to maintain
Cell Lines
Definition:
Cells that are subcultured from primary cells and
adapted to grow in laboratory conditions.
Types
1. Finite cell lines
● Limited number of divisions (20–80 generations)
● Example: Human fibroblasts
2. Continuous cell lines
● Unlimited growth due to mutation or
transformation
● Immortal cells
Examples
Cell line Origin Application
HeLa Human cervical cancer Cancer research
CHO Chinese hamster ovary Protein production
Vero Monkey kidney Vaccine production
3. Maintenance of Cell Culture
Requirements
● Sterile environment (Laminar airflow hood)
● Proper temperature (37°C)
● Proper pH
● Nutrient supply
Subculturing (Passaging)
Definition: Transfer of cells from old medium to fresh medium.
Steps
1. Remove old medium
2. Wash with PBS
3. Add trypsin enzyme
4. Detach cells
5. Transfer to new medium
4. Culture Media and Components
Definition
Culture medium provides nutrients required for cell growth.
Components and Functions
Component Function
Amino acids Protein synthesis
Glucose Energy source
Vitamins Enzyme cofactors
Salts Osmotic balance
Serum Growth factors
Examples of Media
● DMEM (Dulbecco’s Modified Eagle Medium)
● MEM (Minimum Essential Medium)
● RPMI-1640
5. Anchorage Dependence
Definition:
Cells require attachment to surface for growth.
Reason
● Attachment activates growth signals
● Prevents apoptosis
Examples
● Fibroblasts
● Epithelial cells
6. Contact Inhibition
Definition:
Normal cells stop dividing when they contact neighboring cells.
Importance
● Prevents overcrowding
Cancer cells
● Lack contact inhibition
● Grow uncontrollably
7. Types of Animal Cell Culture
1. Organ Culture
Whole organ maintained in culture.
Example: Kidney organ culture
Advantages
● Maintains structure
● Maintains function
2. Explant Culture
Small tissue fragment cultured.
Cells migrate out of tissue.
3. Primary Culture
First culture from tissue.
4. Secondary Culture
Subculture of primary culture.
5. Established Cell Lines
Immortal cells capable of unlimited growth.
Example: HeLa cells
8. Growth Kinetics of Animal Cells
Growth Curve Phases
1. Lag Phase
● Cells adapt
● No division
2. Log Phase
● Rapid division
● Maximum growth
3. Stationary Phase
● Growth rate = death rate
4. Death Phase
● Cells die
9. 3D Cell Culture
Definition
Cells grow in three-dimensional structure.
Advantages
● Mimics natural tissue
● More accurate drug testing
Applications
● Cancer research
● Tissue engineering
10. Transformation of Primary Cells
Definition
Conversion of normal cells into immortal cells.
Methods
1. Viral transformation
Example: SV40 virus
2. Chemical transformation
Carcinogens
3. Radiation
UV radiation
11. Choice of Animal Cells for Protein Production
Most common: CHO cells
Reasons
● Fast growth
● Proper protein folding
● Post-translational modification
Examples of proteins produced
● Insulin
● Growth hormone
● Monoclonal antibodies
12. Viral Vaccine Production
Animal cells used to grow viruses.
Examples
Vaccine Cell line
Polio Vero cells
Rabies BHK cells
COVID-19 Vero cells
13. Scale-Up of Animal Cell Culture
Definition
Increasing culture volume for industrial production.
Methods
1. Spinner flask
Small-scale production
2. Bioreactor
Large-scale production
Types of bioreactors
● Stirred tank bioreactor
● Airlift bioreactor
14. Applications of Animal Cell Culture
Medical applications
● Vaccine production
● Cancer research
● Drug testing
Industrial applications
● Protein production
● Antibody production
Research applications
● Genetic studies
● Toxicity testing
UNIT 2: GENE TRANSFER AND
MANIPULATION METHODS IN ANIMALS
1. Gene Transfer
Definition
Introduction of foreign DNA into animal cells.
Purpose
● Study gene function
● Produce proteins
2. Transformation, Transfection, and Electroporation
Transformation
DNA uptake by cells (common in bacteria).
Transfection
Definition
Introduction of DNA into animal cells.
Methods
Chemical methods
● Calcium phosphate
● Lipofection
Physical methods
● Electroporation
Viral methods
● Retrovirus
● Adenovirus
Electroporation
Definition
Electric pulse creates pores in cell membrane.
Steps
1. Apply electric pulse
2. Membrane pores form
3. DNA enters cell
4. Membrane reseals
3. Stable Transfection
Definition
Foreign DNA integrates into genome permanently.
Selection markers
● Antibiotic resistance gene
Example
● Neomycin resistance
4. Gene Knockdown
Definition
Reducing gene expression.
Methods
RNA interference (RNAi)
Types
• siRNA
• shRNA
Mechanism
1. RNA binds mRNA
2. mRNA destroyed
3. Protein not produced
5. Embryonic Stem Cells
Definition
Pluripotent cells from embryo.
Properties
● Self-renewal
● Differentiate into all cell types
Applications
● Gene therapy
● Transgenic animals
6. Transgenic Animals
Definition
Animals with foreign gene inserted.
Methods
● Microinjection
● Viral vectors
Applications
● Disease research
● Protein production
Example
● Transgenic mice
7. Knockout Animals
Definition
Animals with gene removed.
Purpose
Study gene function.
Example
Knockout mice
8. Cre-LoxP System
Definition
Site-specific recombination system.
Components
● Cre enzyme
● LoxP sites
Function
Removes specific gene
9. Cloning of Animals
Definition
Production of genetically identical animal.
Method: Somatic Cell Nuclear Transfer (SCNT)
Steps
1. Remove nucleus from egg
2. Insert nucleus from body cell
3. Stimulate division
4. Implant into surrogate mother
Example
Dolly sheep
10. Applications of Gene Transfer
● Gene therapy
● Protein production
● Disease research
● Transgenic animals