Part 2:Understanding Plant

Growth In Vitro

Richard R Williams

1. Fundamental Concepts / History

2. Types of Cells or Tissues

3. Cell to Cell Interaction

4. Growth Processes

Topics to be covered:

“… the aseptic culture of plant protoplasts, cells, tissues or organs under conditions which lead to cell multiplication or regeneration of organs or whole plants “

What is Plant Tissue Culture?

Cell theory and totipotency

Callus formation and growth substances

Early cell culture

First plant tissue cultures

Animal tissue cultures

Plant organ culture

1 History

Cork tissue as observed by Robert Hooke in 1664

Cell theory ….

Totipotency ….

… each living cell has a complete genetic blueprint and therefore has the potential to develop into an entire plant.

… cells lines differentiate to form specialised tissues and organs

… unlike animal cells, living plant cells can de-differentiate and then re-differentiate to form different cell types

… complete genetic blue print

… cells differentiate

… living plant cells can re-differentiate

Callus Formation ….…

…Haberlandt .. early 1900’s … proposed concept of totipotency … cells cultured under right conditions

Callus cultured from tree cambium (Gautheret, Nobecourt, Whire in the 1930s.

… cells kept alive but did not develop

Early Cell Culture ….

Cell enlargement … role of auxins

Cell division ... role of cytokinins

- dependent on discovery of “growth regulators”

Early tissue culture ….

Regeneration from tobacco pith ..

(Skoog and Miller) … interaction of auxin

and cytokinin gives differentiation.

GA for growth of shoots

Aux + Cyt + sucrose > vascular development

Culture of ‘thin layers’ … many interacting factors eg pH

Further development …

Carrot plants from root cells – Stewart in 1964

[Steeves & Sussex 1972]

Tobacco plants from single cells – Vasil & Hilderbrandt 1965

[Steeves & Sussex 1972]

Plant Organ Culture ….

Murashige and Skoog 1962 - mineral media

micropropagation

Many different types of cells

Varying degrees of specialisation

- Meristematic- Embryonic- Reproductive

2 Types of Cell & Tissues

Shoot ... apical, … axillary

Meristematic tissues ...

leaf trace

procambium

tunicanew leaf

corpus

pithcortex

apical meristem

axillary meristem

Shoot ... apical, … axillary

Leaf

Root

Meristematic tissues ...

Shoot ... apical, … axillary

Leaf

Root

Adventitious

Cambial tissues

Meristematic tissues ...

Embryonic tissues ...

Pre-formed plant

Generative / somatic

Juvenile

Reproductive tissues ...

Diploid / Haploid

Female / Male

Monocots versus Dicots

‘Stems’ are leaf bases … not buds

Shoot apex at base

Intercallary leaf growth

Important in intact plant

Isolated cell not function the same

Establishment of polarity

Expression of somaclonal (epigenetic)

variation

3 Cell to Cell Interaction

Multiplication Apical dominance Differentiation

Growth Cell division Cell enlargement

Phasic development Juvenility Dormancy

4 Growth Processes

Photosynthesis

Transpiration and water uptake

Cytological instability

4 Other Processes

Physiological mechanisms Physical manipulation Chemical Plant Growth Regulators

(PGRs)

Environment

Genotype

5 Controlling Growth & Development

Hormone theory Auxins Cytokinins

Gibberelins (GA) Ethylene Abscisic Acid (ABA)

Other PGRs

6 Growth Regulators

Hormone BalanceAuxin Cytokinin

High Low

Low High

Root formation on cuttings Embryogenesis

Adventitious root formation in callus Callus initiation

Adventitious shoot formation Axillary shoot growth

Auxin : Cytokinin ratio

Hormone Action …..

Application + uptake

Endogenous + applied

Accumulation & Habituation

Interactions / Sequence

Pulsing vs prolonged exposure

END of Part 2