Conjugation:

One bacterium transfers genetic material to another through direct contact.

Transduction:

A process by which foreign DNA is introduced into a cell by a Virus or viral vector.

Transformation:

A process of gene transfer by which some bacteria take up foreign DNA from environment.

Transfection:

It is a process of introducing naked DNA or RNA into eukaryotic cells.

Cell line:

Cell line is a permanently established all culture that will proliferate indefinitely given appropriate
fresh medium and space.

Cell line development:

1. Tranfection:
Transfect host cells with recombinant plasmids encoding protein of interest.
2. Selection of Pool of transfected cells select cells that are stable and Producing protein of interest.
3. Single cell isolation and all viability single viable cells must be isolated I and cloned in order to
ensure that cell population.
4. Monoclonal assurance-
when developing cell line for biotherapeutics - it is crucial' from Quality and regulatory Pru
spective to ensure that the cell line.
5. Clonal Screening is selection screen and select clones for high expression of protein of interest.

6. Cell Line characterization validate and characterize each clone for stability and productivity.

7. Expansion and downstream evaluation expand clones and perform downstream bioprocesses.
Cell banking is also performed.

Advantages all culture:

1. Scale up is possible.
2. Development of cell line over many generations

Disadvantages of cell culture:

1. Cells may loose some differentiated Characteristics.

 Difference between Batch and continuous culture:

Batch culture Continuous culture

An open system. A closed system.

Small fermenters are used. Large fermenters are used.

Chance of contamination is more. Chance of contamination is less.

Yield is high. Yield is low.

Lag and log Phases are maintained. Lag, log and stationary phases оссur.

Nutrients are continuously added throughout the Nutrients are added at the beginning
process. Process.
Difference between plasmid DNA and chromosomal DNA:

Plasmid DNA Chromosomal

Found in Bacteria only. Found in all the cells.

Circular DNA. Can be circular or linear DNA.

Used as a vector. Not used as a vector.

Replicates independently. Replicates with genome.

Extra- nuclear DNA. Nuclear DNA.

Found in cytoplasm. Found in nucleus.

Always double stranded. May be single stranded or double Stranded.

Naked without histone proteins. Coated with histone proteins.

Does not Carry any vital gene necessary for cell. Carries vital genes necessary for all.
Difference between DNA polymerase and Klenow fragment:

DNA Polymerase Klenow Fragment

A type of enzyme that is responsible for new copies of A large protein fragments of E. coli DNA Polymerase
DNA. I.

Catalyzes the synthesis of DNA molecules. Enzymatic cleavage of Protease subtilisin.

Contain Contain only

5’-3’ Polymerase, 5’-3’ Polymerase,
3’-5’ Exo nuclease, 3’-5’ Exo nuclease.
5’-3’ Exonuclease activity
Synthesizes DNA molecules. Remove 3' and fill-in of 5' overchange to form blunt
ends.

Molecular weight 109 kDa. Molecular weight 68 kDa.

Optimal temp- 72°C. Optimal temp - 25°C.