Lec.
5 Biotechnology
Plant tissue culture
Plant tissue culture: is the growth of explant (any plant part) or plant cells in vitro (in the
laboratory culture media).
• Plant cell culture based on the unique property of the cell-totipotency.
• Cell-totipotency is the ability of the plant cell to regenerate into a whole plant under
the laboratory conditions using artificial nutrient mediums.
• Nutrient media
The nutrient media vary in type and quantity of materials used and the choice
depends on the type of plant to be grown. Some have a better growth in solid media
than in liquid media. The best known nutrient media are: White’s medium, MS
medium, B5 medium. The ideal pH of the culture medium is 5.8, and if the pH values
of more than 7 or less than 4.5, there is inhibition of growth.
Stages of plant tissue culture
There are four stages of plant tissue culture:
1. Initiation stage. A piece of plant tissue (called an Explant) is (a) cut from the plant,
(b) disinfected (removal of surface contaminants), and (c) placed on a medium. A
medium typically contains mineral salts, vitamins, sucrose, antibiotics (optional), and
a solidifying agent such as agar. The objective of this stage is to achieve an aseptic
culture. An aseptic culture is one without contaminating bacteria or fungi.
2. Multiplication stage. A growing of explant can be induced to produce vegetative
shoots by including a cytokinin in the medium. A cytokinin is a plant growth regulator
that promotes shoot formation of growing plant cells.
3. Rooting stage. Growing shoots can be induced to produce adventitious roots by
including an auxin in the medium. Auxins are plant growth regulators that promote
root formation.
4. Acclimatization. A growing, rooted shoot can be removed from tissue culture and
placed in soil. When this is done, the humidity must be gradually reduced over time
because tissue-cultured plants are extremely susceptible to wilting.
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Stages of plant tissue culture
Types of cultures
• Organ Culture (such as the ovary culture and endosperm culture).
• Explant culture (stem, root, flower.... etc).
• Callus culture (is a mass of undifferentiated parenchyma cells).
• Cell suspension cultures.
• Protoplast culture (The protoplast is a cell without cell wall and it obtained by using
enzymes such as cellulases and pectinases).
• Embryo culture.
• Anther and Pollen Culture (They used in the production of haploid plants).
Cell suspension cultures
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Some Applications of Cell and Tissue Culture
1- Micropropagation /Clonal Propagation
• Clonal propagation is the process of a sexual reproduction by multiplication of
genetically identical copies of individual plants.
• Micropropagation is the tissue culture methods of plant propagation. The
micropropagation is rapid and has been adapted for commercialization of important
plants such as banana, apple, and other plants.
2- Production of virus free plants
It has become possible to produce virus free plants through tissue culture at the
commercial level. Among the culture techniques, meristem-tip culture is the most reliable
method for virus and other pathogen elimination.
3- Production of synthetic seeds
In synthetic seeds, the somatic embryos are encapsulated in a suitable matrix (e.g.
sodium alginate), along with substances like mycorrhizae, insecticides, fungicides and
herbicides.
4- Production of secondary metabolites
The most important chemicals produced using cell culture are secondary metabolites.
These secondary metabolites include alkaloids, glycosides (steroids and phenolics),
terpenoids, latex, tannins etc.
Transgenic plants with beneficial traits
1- Transgenic plants or transgenic crops are the plants, in which a functional foreign gene
has been incorporated by any biotechnological methods that generally are not present in the
plant.
2- Transgenic plants have many beneficial traits like insect resistance, herbicide tolerance,
delayed fruit ripening, improved oil quality, weed control etc, but the main goal of producing
transgenic plants is to increase the productivity.
Some of the traits introduced in these transgenic plants are as follows:
Stress tolerance
Biotic stresses (viral, bacterial infections, pests and weeds) and abiotic stresses (physical
actors such as temperature, humidity, salinity etc).
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Herbicide tolerance
Several biotechnological strategies for weed control are being used e.g. the over-
production of herbicide target enzymes (usually in the chloroplast) in the plant which makes
the plant insensitive to the herbicide.
Virus resistance
There are several strategies for engineering plants for viral resistance, and these utilizes
the genes from virus itself. The virus coat protein-mediated approach is the most successful
one to provide virus resistance to plants.
Insect resistance
The transgenic technology uses eco-friendly method to improve pest control
management. The first genes available for genetic engineering of crop plants for pest
resistance were Cry genes (popularly known as Bt genes) from Bacillus thuringiensis. These
are specific to a particular group of insect pests, and are not harmful to other useful insects
like butterflies and silk worms.
Delayed fruit ripening
The gas hormone, ethylene regulates the ripening of fruits, therefore, ripening can be
slowed down by blocking or reducing ethylene production. This can be achieved by
introducing ethylene forming gene(s) in a way that will suppress its own expression in the
crop plant.
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Some of the uses of transgenic plants are:
• Improvement of Nutrient quality
• Improvement of seed protein quality
• Diagnostic and therapeutic proteins
• Edible vaccines
• Biodegradable plastics
