lec_4_nu

ABT 401/Agrl Biotech/NM Boopathi/Lec 4 Nutritional Requirements

Tissue culture media

Nutritional requirements for

optimal in vitro growth of a plant

tissue may vary with the species.

Virtually all plant tissue culture

media are having inorganic

minerals and complex organics and

other nutrients. These include,

1. Inorganic nutrients

2. Carbon / Energy source

3. Vitamins

4. Plant growth regulators

5. Solidifying agent

6. Amino acids

7. Undefined supplements,

and

jasmonates

Role of media constituents

1. Mineral elements are very

important in the life of a

plant. These are constituents

of the cellular organelles

(e.g. magnesium is a part of

chlorophyll molecule). In

addition to C, H and O,

Macro (N, P, K, Ca, S, Mg)

and micro (Fe, Zn, Mn, Cu,

Bo, Mo) elements are the

basic nutrients present in all

the media. They are

essential for normal cell

growth and development.

2. Carbon source (e.g. sucrose)

in the medium is necessary

for various metabolic

activities and act as major

osmotic component in the

medium.

3. For optimum callus growth,

vitamins are required. For

example, thiamine is

essential for proper callus

induction and inositol is

essential in promoting the

callus induction.

4. Plant growth regulators have

great role in tissue culture.

Auxins (e.g. IAA, NAA, 2, 4-

D) are commonly used to

support cell division, callus

growth, somatic embryo

induction, rooting etc.,

Cytokinins (e.g. Kinetin,

Zeatin) are employed to

promote cell division,

regeneration of shoots and

to enhance proliferation and

growth of auxiliary buds.

Abscisic acid (ABA) promotes

somatic embryo and shoot

bud regeneration and

somatic embryo maturation.

Gibberellins (GA3) promotes

shoot elongation and

somatic embryo

germination.


5. In static cultures, if liquid

medium is used the tissue

would get submerged and

die due to lack of oxygen. A

gelling agent (e. g. Agar,

Agarose) is generally used to

circumvent this problem.

6. Complex additives such as

yeast extract, coconut milk,

casein hydrolysate, corn

milk, tomato juice were used

to promote the growth of

certain calli and organs. But

these additives are now no

more preferred, because

their effects in the media

could be reproduced by a

single amino acid such as L-

asparagine or L-glutamine.

Physical form of media:

I. Solidified Medium:

Advantages:

1. Explants are easily seen and

recovered.

2. No Special aeration

required.

3. Shoots grow more orderly.

4. Long term maintenance is

possible.

5. Use of simple containers

occupy little space

Disadvantages:

1. Slower rate of multiplication.

2. Limited surface of explant is

in contact with the medium.

II. Liquid medium:

Advantages:

1. Callus easily break up and

shed as cells to establish a

fine suspension.

2. Faster rate of multiplication

3. A greater surface of explants

is in contact with medium.

4. Toxic metabolites will

effectively be dispersed.

Disadvantages :

1. Recovery is difficult.

2. Growth is disoriented.

3. Seeds would not germinate.

4. Protocorms and plantlets

become brown and dry.

Seeds submerged will show

vitrification.

Commonly used tissue culture

media:

1. MS (Murashige and Skoog,

1962) and LS (Linsmaier and

Skoog, 1972) media are

used for regeneration of

both monocots and dicots.

2. B5 (Gamborg et al., 1969)

developed for culture of

soybean cell suspensions but

also has been effectively

used for variety of plant

regeneration. B5 and its


various derivatives have

been valuable for cell and

protoplast cultures.

3. SH: Schenk and Hildebrandt

(1972) introduced this for

culture of monocots and

dicots. Widely used for

legumes.

4. WPM: Lloyd and McCown

(1980-1981). This is post MS

media. WPM is increasingly

used for propagation of

Ornamental shrubs and trees

in commercial labs.

5. N6 was developed by Chu

for cereal anther culture and

also used for anther culture

of other species.

6. In special cases , NN (Nitsch

and Nitsch) was also used.

Environmental factors

influencing plant tissue

culture:

1. Genotype or variety of the

plant material

2. Explant selection and its size.

3. Medium:

(i) Nutrients,

(ii) Growth regulators and

(iii) Other additives

4. Culture Environment:

(i) Temperature,

(ii) Relative Humidity (RH)

and

(iii) Light

Do you know….

1. What will happen if you alter

the pH of the media?

2. What will be the

consequence if you

autoclave the thermolabile

compounds present in the

media?

lec_4_nu

Documents