Content
Introduction
The practice of plant tissue culture has changed the way some nurserymen
approach plant propagation. In the recent past, the applicability of this technology
to the propagation of trees and shrubs has been documented. Some firms have
established tissue culture facilities and commercial scale operations are presently
in operation for the mass propagation of apples, crabapples, rhododendrons,
and a few other selected woody species. The intent of this research update is
to briefly examine “what is being done” and to explore “what can be done”
with regard to the tissue culture of ornamental plants. Such a consideration
necessarily includes an overview of tissue culture as a propagation tool. The
major impact of plant tissue culture will not be felt in the area of
micropropagation, however, but in the area of controlled manipulations of plants
at the cellular level, in ways which have not been possible prior to the
introduction of tissue culture techniques.
The Art and Science of Micropropagation
“Micropropagation” is the term that best conveys the message of the tissue
culture technique most widely in use today. The prefix “micro” generally refers
to the small size of the tissue taken for propagation, but could also refer to the
size of the plants which are produced as a result.
Micropropagation allows the production of large numbers of plants from
small pieces of the stock plant in relatively short periods of time. Depending on
the species in question, the original tissue piece 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 plant is placed in tissue culture, proliferation
of lateral buds and adventitious shoots or the differentiation of shoots directly
from the callus, results in tremendous increases in the number of shoots available
for rooting. Rooted “microcuttings” or “plantlets” of many species have been
established in production situations and have been successfully grown on
either in containers or in field plantings. The 2 most important lessons learned
from these trials are that this methodology is a means of accelerated asexual
propagation and that plants produced by these techniques respond similarly to
any own-rooted vegetatively propagated plant.
Micropropagation offers several distinct advantages not possible with
conventional propagation techniques. A single explant can be multiplied into
several thousand plants in less than 1 year. With most species, the taking of
the original tissue explant does not destroy the parent plant. 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.
Plant Improvement Through Tissue Culture
In introducing this research update, it was mentioned that the major impact of
tissue culture technology would not be in the area of micropropagation, but
rather in the area of controlled manipulations of plant germplasm at the cellular
level. The ability to unorganize, rearrange, and reorganize the constituents of
higher plants has been demonstrated with a few model systems to date, but
such basic research is already being conducted on ornamental trees and shrubs,
with the intent of obtaining new and better landscape plants.
Selection of Plants with Enhanced Stress or Pest Resistance
Perhaps the most heavily researched area of tissue culture today is the concept
of selecting disease-, insect-, or stress-resistant plants through tissue culture.
Just as 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.
All who are familiar with bud sports, variegated foliage, and other types
of chimeras have an appreciation for the natural variability in the genetic
makeup or expression in plants. Chimeras are the altered cellular expressions
that are visible, but for each of these that are observed many more differences
probably exist but are masked by the overall organization of the plant as a
whole. For example, even in frost-tender species, certain cells or groups of cells
may be frost-hardy. However, because most of the organism is killed by frost,
the tolerant cells eventually die because they are unable to support themselves
without the remainder of the organized plant.
Plant tissues grown in vitro can be released from the organization of the
whole plant through callus formation. If these groups of cells are then subjected
to a selection agent such as freezing, then those tolerant ones can survive while
all those that are susceptible will be killed. This concept can be applied to many
types of stress, as well as resistance to fungal and bacterial pathogens and
various types of phytotoxic chemical agents. Current research in this area extends
across many interests, including attempts to select salt-tolerant lines of tomato,
freezing-resistant tobacco plants, herbicide-resistant agronomic crops, and various
species of plants with enhanced pathogen resistance. Imagine, if you will, the
impact of a fireblight-resistant Bartlett pear, a clone of pin oak for alkaline soils,
or a selection of southern magnolia hardy to zone 4.
Tissue Culture and Pathogen-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.
The concept behind indexing plants free of pests is closely allied to the concept
of using tissue culture as a selection system. Plant tissues known to be free of
the pathogen under consideration (viral, bacterial, or fungal) are physically
selected as the explant for tissue culture. In most cases, the apical domes of
rapidly elongating shoot tips are chosen. These are allowed to enlarge and
proliferate under the sterile conditions of the in vitro culture with the resulting
plantlets tested for presence of the pathogen (a procedure called indexing).
Cultures that reveal the presence of the pathogen are destroyed, while those that
are indexed free of pathogen are maintained as a stock of pathogen-free material.
Procedures similar to these have been used successfully to obtain virus-free
plants of a number of species and bacteria-free plants of species known to have
certain leaf-spot diseases. The impact of obtaining pathogen-free nursery stock
can only be speculative, since little research documenting viral, bacterial, or
fungal diseases transmitted through propagation of woody ornamentals is
available.
Somatic Hybridization
The ability to fuse plant cells from species that may be incompatible as sexual
crosses and the ability of plant cells to take up and incorporate foreign genetic
codes extend the realm of plant modifications through tissue culture to the
limits of the imagination. Most such manipulations are carried out using plant
“protoplasts”. Protoplasts are single cells that have been stripped of their cell
walls by enzymatic treatment. A single leaf treated under these conditions may
yield tens of millions of single cells, each theoretically capable of eventually
producing a whole plant. This concept has fueled speculation as diverse as the
possibilities of obtaining nitrogen-fixing corn plants at one extreme, to discovering
a yellow-flowered African violet at the other extreme.
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. It is conceivable then that one could hybridize a juneberry with a
crabapple or a plum, but the fundamental research required to demonstrate
such an event has yet to be conducted.
Summary
Plant tissue culture research is multidimensional. While most nurserymen have
been introduced to the techniques and advantages of micropropagation, few
have ventured to use it as a propagation tool. The applicability of micropropagation
for woody trees has been demonstrated as feasible, since all aspects of
the technology have confirmed the fact that trees produced by this method look
like and grow like their counterparts produced by traditional methods of cloning.
Other dimensions of tissue culture research have been less well publicized.
The potential for selecting pathogen-free plants, for selecting stress-tolerant and
pathogen-resistant clones of plants, and the novel genetic combinations to be
achieved through somatic hybridization are all lines of research that can have
a profound impact on the nursery industry.
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