Lecture 1

By

Dr. Ahmed Metwaly

TISSUE CULTURE

Objectives:■ Definition

■ History

■ Some basics

■ Factors affecting tissue culture

■ Types of tissue culture

■ Tissue culture and agriculture

■ Tissue culture and pharmacy

■ Factors necessitate the development of tissue culture

Definition:

■ Tissue culture is the in vitro aseptic culture of cells, tissues, organs or

whole plant under controlled nutritional and environmental conditions

History of plant tissue culture:1838-39 cellular theory (Cell is totipotent) Schleiden-

Schwann

1902 First attempt of plant tissue culture Haberlandt

1939 Continuously growing callus culture White

1946 Whole plant developed from shoot tip Ball

1950 Organs regenerated on callus Ball

1954 Plant from single cell Muir

1960 Protoplast isolation Cocking

1962 MS media Murashige -

Skoog

1964 Clonal propagation of orchids Morel

1964 Haploids from pollen Guha

1970 Fusion of protoplasts Power

1971 Plants from protoplasts Takebe

1981 Somaclonal variation Larkin

Haberlandt Earnest A. Ball

Some Basics;

The Architecture of Plants

TotipotencyThe total potential of a plant cell to develop into an entire plant if

suitably stimulated.

DedifferentiationCapacity of mature cells to return to meristematic condition and

development of a new growing point, follow by redifferentiation which is the ability to reorganize into new organ.

ExplantThe tissue obtained from a plant to be cultured.

Callus A proliferating mass of undifferentiated plant parenchyma cells

derived from plant tissue (explants) for use in biological research.

This type of cells can be produced by plants as a result of a wound.

Callus

Clone

A group of plants propagated only by vegetative or asexual means, all members

of which have been derived from a single individual.

Organogenesis

A process of differentiation by which plant organs are formed from tissue or

callus or de novo differentiation of organs as separate entities, i.e., roots and shoots.

Somatic embryogenesis

An in vitro plant regeneration process from somatic cells that involves

differentiation via a somatic embryo which mimics a zygotic embryo.

Protoplast

A single plant cell from which the cell wall has been removed (usually by use cell

wall degrading enzymes).

Protoplast

Factors affecting tissue culture

■ Growth Media

– Minerals, Growth factors, Carbon source, Hormones (Two Hormones Affect Plant Differentiation):

– Auxin: Stimulates Root Development

– Cytokinin: Stimulates Shoot Development

– Auxin ↓Cytokinin = Root Development

– Cytokinin ↓Auxin = Shoot Development

– Auxin = Cytokinin = Callus Development

■ Environmental Factors

– Light, Temperature, Photoperiod, Sterility.

– Explant Source

Usually, the younger, less differentiated the explant, the better for tissue culture

Types of In vitro tissue culture

■ Culture of intact plants (Clonal)

■ Embryo culture (Somatic embryogenesis )

■ Organ culture

1. shoot tip culture

2. root culture

3. leaf culture

4. anther culture

■ Callus culture

■ Cell suspension culture

■ Protoplast culture

Tissue culture in agriculture

■ Production of improved crop varieties

■ Production of disease-free plants (virus)

■ Genetic transformation

■ Production of varieties tolerant to salinity, drought and heat

stresses

Tissue culture in pharmacy

■ The elucidation of the biosynthetic pathways of secondary metabolites

with isolation of corresponding enzymes.

■ Discovery of new secondary metabolites in vitro.

■ The commercial production of expensive secondary metabolites .

■ They have also been used for metabolic and genetic studies.

■ The selection of superior strains of medicinal plants.

Factors necessitate the development of tissue culture

1. Availability of raw material:

Some plants cannot be produced in economically sufficient

quantity to satisfy demand, e.g. Taxus species, the principal

source of a diterpene alkaloid; taxol.

2. Variation and fluctuation of supplies and quality:

The production as well as the quality of crude drugs is affected by

climatic variability, crop diseases and various methods of

collection and drying. In addition, variation in the active

constituents may arise in plants of the same species having

different genetical characteristics

. On contrary, plant tissue culture techniques would permit a steady growth of

tissues or cells away from the effect of the above variables.

• Growing plant cells or tissues under optimum environmental conditions, i.e.

better control can be attained of light, temperature and nutrition.

• Production of active constituents could be continuous at all times and at a

standard quality.

• Growing plant cells or tissues yield no artifacts or stress metabolites attributed

to bacteria, fungi, algae, viruses and insects.

Summary:■ Definition

■ History

■ Some basics

■ Factors affecting tissue culture

■ Types of tissue culture

■ Tissue culture and agriculture

■ Tissue culture and pharmacy

■ Factors necessitate the development of tissue culture