CALLUS CULTURE

Agustina Monalisa Tangapo

BRAINSTORMING

Characteristic of plant in
vitro culture
1. Environmental condition optimized (nutrition,
light, temperature).
2. Ability to give rise to callus, embryos,
adventitious roots and shoots.
3. Ability to grow as single cells (protoplasts,
microspores, suspension cultures).
4. Plant cells are totipotent, able to regenerate a
whole plant.


Three fundamental abilities of
plants
Totipotency

Dedifferentiation

Competency



Important factors
Growth Media
Minerals, growth factors, carbon source, hormones
Environmental Factors
Light, temperature, photoperiod, sterility, growth media
Explant Source
Usually, the younger, less differentiated explant, the better for
tissue culture
Different species show differences in amenability to tissue
culture
In many cases, different genotypes within a species will have
variable responses to tissue culture; response to somatic
embryogenesis has been transferred between melon cultivars
through sexual hybridization



Hormone



Product Name



Function in Plant Tissue Culture

Auxins

Indole-3-Acetic Acid
Indole-3-Butyric Acid
Indole-3-Butyric Acid, Potassium Salt
-Naphthaleneacetic Acid
2,4-Dichlorophenoxyacetic Acid
p-Chlorophenoxyacetic acid
Picloram
Dicamba


Adventitous root formation (high concen)
Adventitious shoot formation (low concen)
Induction of somatic embryos
Cell Division
Callus formation and growth
Inhibition of axillary buds
Inhibition of root elongation

Cytokinins

6-Benzylaminopurine
6-,-Dimethylallylaminopurine (2iP)
Kinetin
Thidiazuron (TDZ)
N-(2-chloro-4-pyridyl)-NPhenylurea
Zeatin
Zeatin Riboside

Adventitious shoot formation
Inhibition of root formation
Promotes cell division
Modulates callus initiation and growth
Stimulation of axillarys bud breaking and growth
Inhibition of shoot elongation
Inhibition of leaf senescence

Gibberellins

Gibberellic Acid

Stimulates shoot elongation
Release seeds, embryos, and apical buds from dormancy
Inhibits adventitious root formation
Paclobutrazol and ancymidol inhibit gibberellin synthesis thus
resulting in shorter shoots, and promoting tuber, corm, and bulb
formation.

Abscisic Acid

Abscisic Acid

Stimulates bulb and tuber formation
Stimulates the maturation of embryos
Promotes the start of dormancy

Polyamines

Putrescine
Spermidine

Promotes adventitious root formation
Promotes somatic embryogenesis
Promotes shoot formation


Callus
An un-organised mass of cells
A tissue that develops in response
to injury caused by physical or chemical means
Most cells of which are differentiated although may be and are
often highly unorganized within the tissue
It can be multiplied through solid or suspension culture
Callus (mass of parenchymatous cells)
organogenesis
Somatic embryogenesis
Plant regeneration
Kalus adalah suatu
kumpulan sel amorphous
yang terjadi dari sel-sel
jaringan yang membelah
diri secara terus menerus
BAGAIMANA MENDAPATKAN
KULTUR KALUS????
Two hormones affect plant differentiation:
Auxin: Stimulates root development
Cytokinin: Stimulates shoot development
Generally, the ratio of these two hormones can
determine plant development:
Auxin Cytokinin = Root development
Cytokinin Auxin = Shoot development
Auxin = Cytokinin = Callus development
Control of in vitro culture
Cytokinin
Auxin
Leaf strip
Adventitious
Shoot
Root
Callus

TUGAS!!!!!!! (^_^)
Callus culture




Explants Callus



Protoplasts

Development
Suspension cells

Organs

(leaves, roots, shoots, flowers,...)

De-differentiation Re-differentiation
1. Meristems

2. Leaf sections

3. Bulb sections

4. Embryos

5. Anthers

6. Nucellus

Callus formation
Stimuli :

In vivo : wound, microorganisms, insect feeding


In vitro : Phytohormones
1. Auxin
2. Cytokinin
3. Auxin and cytokinin
4. Complex natural extracts

Callus formation
Callus
During callus formation there is some degree of
dedifferentiation both in morphology and
metabolism, resulting in the lose the ability to
photosynthesis.
Callus cultures may be compact or friable.
Compact callus shows densely aggregated cells
Friable callus shows loosely associated cells and
the callus becomes soft and breaks apart easily.
Habituation:
The lose of the requirement for auxin and/or
cytokinin by the culture during long-term culture.

Genotypic effects on callus morphology

Three stages of callus culture
Induction: Cells in explant dedifferentiate and begin
to divide
Proliferative Stage: Rapid cell division
Differentiation stage (sometimes): organogenesis
or embryogenesis
Persiapan eksplan


Induction
1998-2003, Branch of Shemyakin&Ovchinnikov IBCh RAS
Division
E. Sutton, UC Davis
Callus
1998-2003, Branch of Shemyakin&Ovchinnikov IBCh RAS
Andrographis paniculata


Kalus yang tumbuh pada eksplan tunas
pucuk manggis

Figure 1. Stages of callus growth in A.
malaccensis Lam.a. Initiation of callus, b&c;
Growth of the callus; d. Cell death
Differentiation
Organogenesis
Somatic embryogenesis

Gambar. Induksi dan proliferasi kalus (1-2);
Diferensiasi/(regenerasi tunas (3-4) dan Pembentukan plantlet
induksi perakaran(5-6)
Lolium temulentum
MANFAATNYA?????
Callus culture of some
commercially important plants

Podophyllum
hexandrum
Azadirachta indica
Linum album
THANKS FOR UR ATT....