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Technology
Review on The Many Applications of Plant
Tissue Culture Research
Introduction
Plant tissue culture can be defined as the culture of plant
cells, tissue and organs under aseptic conditions.
Plant tissue culture has an important role to play in the production
of ornamental or agricultural plants and in the manipulation of
plants for improved agronomic performance. Plant tissue culture
research is a multi-dimensional science that offers exciting prospects
to future improvements in crop productivity. While most nurserymen
have been introduced to the techniques of micropropagation, other
dimensions of tissue culture research have been less well publicized.
For example, the potential for selecting pathogen- or stress-resistant
plant clones, the creation of novel genetic combinations through
somatic hybridization,… are techniques that have been unfrequently
transposed to the nursery industry.
In this article, a rapid overview of some developments which can
have a deep impact on the nursery industry are presented. Many
articles which discuss in detail the basic procedures and methods
involved in plant culture, may be consulted for complementary informations (Pierick
1987; Debergh and Zimmerman 1991).
The area under review include :
- Micropropagation.
- Meristem culture and production of disease-free plants.
- Somatic embryogenesis.
- Somaclonal variation.
- In vitro selection.
- Protoplast culture and somatic hybridization.
1. Micropropagation
One of the main application of micropropagation is the mass propagation
of superior plants. In many instances, conventional propagation
is a slow process during which disease and pest problems can limit
production. Micropropagation offers the potential to produce thousands,
or even billions of plants per year. Micropropagation offers several
advantages not possible with conventional propagation techniques.
Once established, actively dividing cultures are a continuous source
of microcuttings which can result in plant production under greenhouse
conditions, without seasonal interruption. Using methods of micropropagation,
the nurseryman can rapidly introduce selected superior clones of
ornamental plants in sufficient quantities to have an impact on
the landscape plant market.
Micropropagation allows the production of large numbers of plants
from small pieces of a stock plant in relatively short periods
of time.
Depending on the species, the original tissue explant may be taken
from shoot tip, leaf, lateral bud, stem or root tissue. In most
cases, the original plant is not destroyed in the process, a factor
of considerable importance to the owner of a rare or unusual plant.
Once the explant is placed on an appropriate culture medium, proliferation
of buds and adventitious shoots results in tremendous increases
in the number of shoots. Subculture of the buds or shoot is repeated
until many plants are produced, all having the genetic characteristics
of the original specimen.
The mean proliferation rate depends on the species treated. Shoots
are generally separated every four weeks and transferred onto a
fresh proliferation medium. Induction of the root system on indivifual
shoot may be induced on appropriate medium.
Rooted "microcuttings" or "plantlets" of many
species have been established in production situations and have
been successfully grown either in containers or in field plantings.
2. Meristem culture and production of disease-free
plants
Another purpose for which plant tissue culture is uniquely suited
is in the obtaining, maintaining, and mass propagating of specific
pathogen-free plants by meristem culture.
Meristem culture was pioneered by Morel (1960) and
usually involves the removal of the meristem and subsequent culture
on a nutrient medium. The meristem is a dome of activelly dividing
cells, about 0.1 mm in diameter. Endogeneous contaminants do not
easily invade in the meristem, often resulting in the formation
of a disease-free plant. When combined with micropropagation techniques,
large numbers of disease-free plants may be produced from meristematic
explants.
Meristem culture has been used successfully in the removal of viruses
from many plants (potato, sugarcane, strawberry) (Quak, 1977) and
is now used routinely for the eradication of many viral diseases
from plant material.
3. Somatic embryogenesis
Somatic embryogenesis refers to the development of embryo-like
structures from cells of somatic (non-sexual) origin onto an appropriate
medium. Somatic embryo occurs either directly on the explant or
more often in callus culture.
There are several advantages of recovery of plants from cells via
somatic embryogenesis compared to micropropagation.
- Somatic embryos can be produced from cells growing in suspension,
thereby making possible batch culture techniques which can
be scaled-up with minimum handlings costs.
- The multiplication rate is very high and, in some species
like carrot, celery or tomato, the embryos can be encapsulated
and treated as an artificial seeds.
- Whole plants develop from the somatic embryos and only require
growth to maturity. When organogenesis occurs, shoots or roots
develop and the induction of the complementary structures frequently
requires different culture media formulations.
However, many problems remain with somatic embryogenesis.
- Altough callus proliferation is relatively easy for most
plant species, regeneration from unorganised cells, such as
cell suspensions, is usually a more difficult process.
- Moreover, there are numerous reports that show that the regeneration
from callus or cell-suspensions may lead to genetic variations
in regenerated plants. Therefore, it is important that more
research be conducted to understand the molecular basis of
the genetic changes that occur during the artificial culture
of cells.
4. Somaclonal variation
Although conventional micropropagation has resulted to a large
extent in clonal fidelity, it has become increasingly clear that
under the appropriate culture conditions, a great deal of genetic
variability can be recovered in regenerated plants. If cultures
are established from explants that did not contain a pre-organized
meristem, or if cultures are maintained as callus prior to plant
regeneration, the regenerated plants are quite variable.
In early report, most of the variation were attributed to the readily
detected chromosome instability of cultured plant cells. In many
cases, the degree of instability was reported to be proportional
to the length of time the cells remained in culture.
Recognization of this spontaneous variation inherent in long-term
culture led to the use of cell culture for mutagenesis and selection
of genetic variants and for direct recovery of novel genotypes
from cell cultures via somaclonal variation.
Indications of somaclonal variation in several crop plants have
stimulated interest in application of this method for crop improvement.
There is now several cases where somaclonal variation had produced
agricultural useful changes in the progeny (e.g. eye-spot resistance
or increase in sugar yeld in sugarcane - late blight resistance
in potato - resistance to Fusarium in tomato) (Evans, 1989).
5. In vitro selection
Today, one of the most intensively studied area of tissue culture
is the concept of selecting disease, insect, or stress resistant
plants through tissue culture.
Significant gains in the adaptability of many species have been
obtained by selecting and propagating superior individuals, so
the search for these superior individuals can be tremendously accelerated
using in vitro systems.
Such systems can attempt to exploit the natural variability known
to occur in plants or variability can be induced by chemical or
physical agents known to cause mutations.
In vitro selection usually involves subjecting a population
of cells to a suitable selection pressure and recovering any variant
lines which have developed resistance or tolerance to the stress.
The goal would be to reorganize whole plants from such resistant
cell lines. This approach presumes that tolerance operating at
the unorganised cellular level can act to some degree of effectiveness,
in the whole plant. If the tolerance has a genetic basis then the
trait can be transferred to other plants.
Current research in this area extends across many interests including
attempts to select salt tolerant lines, freezing resistant plants,
herbicide resistant agronomic crops, and various resistance to
chemical molecules such as heavy metal (aluminium, manganese, …).
6. Protoplast culture and somatic hybridization
Protoplasts are single cells which have been stripped of their
cell walls by enzymatic treatment. A single leaf treated under
these conditions can give millions of single cells, each theoretically
capable to produce a whole plant.
The observation, that has provided the impetus for most of this
research, is that when cells are stripped of their cell walls and
brought into close contact, they tend to fuse with each other.
This "somatic hybridization" is not subject to the same
incompatibility problems that limit traditional plant breeding
strategies. The ability to fuse plant cells from species which
may be incompatible as sexual crosses extend the realm of plant
modifications through tissue culture to the limits of the imagination.
The potential use of somatic hybridization to bring about novel
combinations of genetic material had been demonstrated in the genera Petunia and Nicotiana.
Further research in this area promises to have a tremendous impact
on our concept of plant diversity.
References
Debergh
P.C. and Zimmerman R.H. (1991).
Micropropagation : Technology and Application.
Kluwer Academic Publishers.
Evans
D.A. (1989).
Somaclonal variation - genetic basis and breeding applications.
Trends
in Genetics, 5, 46-50.
Morel G.M. (1960).
Producing virus-free cymbidiums.
American Orchid
Society Bulletin, 29, 495-497.
Quak F. (1977).
Meristem culture and virus-free plants.
In " Applied and Fundamental Aspects of Plant Cell, Tissue
and Organ Culture". (Eds J. Reinert and Y.P.S. Bajaj pp 596-615).
Springer-verlag Berlin.
Pierik R.L.M. (1987).
"in vitro Culture of Higher Plants". Martinus Nijhoff Publishers.
Dordrecht. |
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