R. Daniel Lineberger
Professor of horticulture
Texas A&M University. M university
College Station, Texas, 77843
The stationmaster of agricultural school horticulture (commercial scale operation) is currently carrying out mass propagation of apples, begonia, rhododendrons and some other selected woody species. The purpose of this latest research is to briefly examine "what is being done" and to explore "what can be done" about tissue culture of ornamental plants. This consideration necessarily includes an overview of tissue culture as a reproductive tool. However, the main influence of plant tissue culture is not in the field of micropropagation, but in the field of controlling plants at the cell level in a way that was impossible before the introduction of tissue culture technology.
The art and science of micropropagation
Among all the terms used in this process, the word "micropropagation" can best express the information of the most widely used tissue culture technology today. The prefix "micro" usually refers to the small size of the tissue used for reproduction, but it can also refer to the size of the resulting plants.
Micropropagation allows a large number of plants to be produced from small original plants in a relatively short time. Depending on the species in question, the original tissue block can be taken from shoot tips, leaves, lateral buds, stems or root tissues (Figure 1). In most cases, primitive plants will not be destroyed during processing, which is a very important factor for the owners of rare or unusual plants. Once plants are placed in tissue culture, the proliferation of lateral buds and adventitious buds (Figure 2) or direct differentiation of buds from callus (Figure 3) leads to a great increase in the number of buds available for rooting. Many species of rooted "micro-cuttings" or "seedlings" have been established in production and have successfully grown in containers or fields. The two most important lessons learned from these experiments are that this method is a means to accelerate asexual reproduction, and the plants produced by these techniques have similar reactions to any plants that reproduce asexually from their roots. Micro-propagation provides several obvious advantages that traditional propagation techniques cannot provide. An explant can reproduce into thousands of plants in less than a year. For most species, obtaining original tissue explants will not destroy the parent plants. Once established, actively dividing cultures are a continuous source of microdissection, which can lead to plant production under greenhouse conditions without seasonal interruption. Using micropropagation, nursery workers can quickly introduce a sufficient number of selected excellent ornamental plant clones to have an impact on the garden plant market.
Plant improvement through tissue culture
When introducing this latest research, it is mentioned that the main influence of tissue culture technology is not in the field of micropropagation, but in the field of controlling plant germplasm at the cell level. Up to now, some model systems have proved the ability of disordering, rearranging and recombining the components of higher plants, but this basic research has been carried out on ornamental trees and shrubs in order to obtain new and better landscape plants.
Selecting plants with enhanced stress or pest resistance
Perhaps the most studied field of tissue culture today is the concept of selecting disease-resistant, insect-resistant or stress-resistant plants through tissue culture. Just as the adaptability of many species has been significantly improved by selecting and breeding excellent individuals, the study of these excellent individuals can be greatly accelerated by using in vitro systems. Such systems may attempt to take advantage of natural variability known to occur in plants, or variability may be induced by chemical or physical agents known to cause mutations.
All people familiar with bud movement, variegated leaves and other types of chimeras appreciate the natural variability of plant gene composition or expression. Chimera is a visible change in cell expression, but there may be more differences for each observed species, but it is covered by the whole plant tissue. For example, even in cold-tolerant species, some cells or cell groups may be cold-tolerant. However, because most organisms are killed by frost, tolerant cells eventually die, because they can't support themselves without organizing the rest of plants. Plant tissues grown in test tubes can be released from the whole plant tissues through the formation of callus. If these cell groups are subjected to Argan selection test, such as freezing, those tolerant cells can survive and all susceptible cells will be killed. This concept can be applied to many types of stress and resistance to fungal and bacterial pathogens and various types of phytotoxic chemicals. The goal of selecting such resistant cell lines is to recombine the whole plant from them, which will maintain the selected resistance (Figure 4). At present, the research in this field spans many interests, including trying to select salt-tolerant tomato strains, frost-resistant tobacco plants, herbicide-resistant crops and various plants with enhanced pathogen resistance. Imagine, if you like, a fire-resistant Bartley pear, a needle oak clone in alkaline soil, or the influence of a cold-resistant southern magnolia to fourth area!
Tissue culture and pathogen-free plants
Another purpose that plant tissue culture is particularly suitable for is to obtain, maintain and multiply plants without specific pathogens. The concept behind pest-free plant index is closely related to the concept of using tissue culture as a selection system. Physically, plant tissues which are known not to contain the considered pathogens (viruses, bacteria or fungi) are selected as explants for tissue culture. In most cases, the top dome of the rapidly elongated stem tip is selected (Figure 5). They are allowed to expand and proliferate in vitro under aseptic conditions (Figure 6), and the generated plantlets are tested for the presence of pathogens (a procedure called indexing). Cultures exposed to the presence of pathogens are destroyed, while those that are indexed as pathogen-free are kept as a reserve of pathogen-free materials. Methods similar to these have been successfully used to obtain virus-free plants of many species and bacteria-free plants of species known to suffer from certain leaf spot diseases. The impact of obtaining pathogen-free seedlings can only be speculated, because few studies have recorded viral, bacterial or fungal diseases transmitted by woody ornamental plants.
Somatic hybridization
The ability to fuse plant cells from potentially incompatible species as sexual hybridization and the ability of plant cells to absorb and integrate exogenous genetic codes extend the field of plant modification through tissue culture to the limit of imagination. Most of these operations are performed with plant "protoplasts". Protoplast is a single cell whose cell wall has been stripped by enzyme treatment. A leaf treated under this condition may produce tens of millions of single cells, and theoretically each single cell can eventually produce a complete plant. This concept has caused various speculations, on the one hand, the possibility of obtaining nitrogen-fixing corn, on the other hand, the discovery of a yellow-flowered African violet.
The observation that powered most of this research was that when cells were stripped of their cell walls and brought into close contact, they tended to fuse with each other (Figure 7). This kind of "somatic hybridization" will not encounter the incompatibility problem that limits the traditional plant breeding strategy. It is conceivable that one can cross June berries with begonia or plums, but the basic research needed to prove this event has not yet been carried out.
The potential of somatic hybridization to produce new genetic material combinations has been proved in Petunia and Nicotiana. Research partially funded by the Horticulture Research Institute of the University of Wisconsin is investigating the feasibility of applying this technology to woody species. Brent McGown and his colleagues have successfully obtained null cell from tissue cultures of birch and rhododendron, but so far, they have neither obtained plants from single cells nor achieved cell fusion. However, further research in this field is expected to have a great impact on our concept of woody plant diversity. As striking as the idea of fusing plant protoplasts is the idea of incorporating foreign genetic material into the genetic code of plant cells. This transformation has been carried out in the so-called "gene splicing" experiment, in which the information of making insulin is integrated into bacteria. Not only the required information is passed on to the next generation of bacteria, but also bacterial cultures become insulin synthesizers. Plant cells can accept foreign genetic codes, but there is no evidence that this genetic code can be passed on to daughter cells and play the expected role. For example, what if the genetic information of high sugar accumulation is integrated into a sugar maple clone? One can think enough if a serious volume is filled in this category!
abstract
The study of plant tissue culture is multidimensional. Although most nursery owners have understood the technology and advantages of micropropagation, few people take the risk to use it as a breeding tool. The applicability of micropropagation of woody trees has been proved to be feasible, because all aspects of this technology have confirmed the fact that the trees produced by this method look and grow similar to those produced by traditional cloning methods.
Other aspects of tissue culture research are not well known. The potential of selecting pathogen-free plants, the potential of selecting stress-tolerant and pathogen-resistant plant clones, and the new genetic combination realized by somatic hybridization are all research directions that can have a far-reaching impact on nursery industry.