cell tissue &Protoplast Culturing

1. TISSUE, CELL & PROTOPLAST
CULTURES
Minahil Khalid
MP-MMG-12

2. Tissue
Culturing.

3.  Tissue culture
It is defined as “the process of growing cells artificially in the laboratory”.
 Plant Tissue culture
It is defined as a “technique for the production of large numbers of
genetically identical plantlets.

4. 1. Aseptic condition,
2. Aeration.
3. Equipment.
4. Nutrient medium.
5. Appropriate tissue(Explant).
6. Frequent Subculturing.
Requirements:

5. Types of Plant Tissue Culture:

6. Process of Plant Tissue Culture

7. 1) Recalcitrance.
2) Contamination.
3) Phenolic browning.
4) Seasonal variation.
5) Vitrification.
Problems in PTC

8. 1. Juvenile tissue can be selected as an explant and treated with
cytokinin spray.
2. Sterilize the working surfaces with ethanol and use sterile
equipment.
3. Culture bottles are kept in dark condition
4. Culture bottles are subculture frequently to overcome vitrification
How to overcome?

9. Cell culturing:
“An artificial process of growing cells under controlled
conditions using artificial media in order to study the
biological properties of cells or cell lines is referred to as
cell culture”.

10. 1. Appropriate culture media
2. Correct temperature
3. Sterile environment
4. Laminar flow hood
Requirements:

11. Composition of cell culture

12.  Sample collection
 Isolating target cells
 Maintaining cells in a culture
 Choosing cell culture media- essential components
 Preventing contamination
 Cell harvesting
Steps in cell culture:

14. Troubleshooting
Microbial
Contamination.
Laminar Flow Hood
Cross-
Contamination.
Authenticate cell
lines using short-
tandem repeat
profiling(STR)
Problem? How do overcome it?

15. Protoplast culturing:
It refers to the process in which whole plants are
developed from the culture of cells without a cell
wall.

16.  Sterilization of leaves,
 Peeling off the lower epidermis of leaves,
 Pre-enzyme treatment and incubation in enzyme,
 Isolation and cleaning of the protoplasts by filtration and
centrifugation
Major Steps :

17. Steps involved in culture isolation & Protoplast regeneration.

18. Factors Affecting :
1) Plant species &Varieties
2) Plant age & organ
3) Pre-culture conditions
4) Temperature
5) Growth Media

19. Applications :
Protoplast culture can be used to study:
For Somatic Hybridization.
For DNA Transformation.
Ingesting foreign material into the cytoplasm.
Wall synthesis & decomposition.