(Please note that it is required of the learner to have a significant knowledge of genetics before this chapter because we will not be discussing “genetics” in micro details).
The natural world is covered by populations of many different kinds of plants that have evolved over eons of time. We identify these as species, although there are other divisions that will be described in this text. These populations can more or less maintain themselves from generation to generation because of their natural genetic characteristics. If not, they evolve into other variants or become extinct. In agriculture and horticulture, on the other hand, propagators primarily deal with special kinds of plants, which are defined as cultivars (varieties) (9). We buy ‘Thompson Seedless’ grapes and ‘Elberta’ peaches for our table, grow ‘Queen Elizabeth’ roses and ‘Bradford’ pear trees in our landscape, and plant ‘Hybrid Yellow Granex’ onion seed and ‘Marquis’ wheat in our fields. All of these represent populations of plants that are unique and only exist in cultivation. These plants would likely change drastically, or disappear altogether, if not maintained by genetic selection during propagation. Chapters 19, 20, and 21 describe the range of cultivated plants and their propagation.
Plant propagation and plant breeding both involve genetic selection. The role of the plant breeder is to recreate patterns of genetic variation in its many forms from which to select new kinds of plants useful to humans. The role of the plant propagator, on the other hand, is to multiply these selected cultivars and to do it in such a manner as to maintain the genetic characteristics of the original population. To do both requires an understanding of genetic principles and procedures.
BIOLOGICAL LIFE CYCLES IN PLANTS
Plant Life Cycles
In natural systems, plant life cycles can be described based on their life span and reproductive pattern. Therefore, they are referred to as annuals, biennials, or perennials:
- Annuals are plants that complete the entire sequence from germination to seed dissemination and death in one growing season. Technically, annuals are monocarpic, meaning that they die after reproducing. However, “annuals” also refers to plants that may be perennial in mild climates but are not winter hardy, and so die after the first growing season due to cold temperatures.
- Biennials are plants that require two growing seasons to complete their life cycle. During the first year, the plants are vegetative and grow as low clumps or a rosette of leaves. These plants usually need a period of cold weather for vernalization of the shoot meristem before they can become reproductive. During the second season, biennial plants bolt, producing a fast-growing flowering spike, flower, produce seeds, and then die. Although the terminology is confusing, winter annuals fit into this category. Seeds germinate in late summer, forming a seedling with numerous rosette leaves that hug the ground. After winter vernalization, the meristem bolts, flowers, sets seeds, and dies before summer (less than 12 months).
- Perennials are plants that live for more than 2 years and repeat the vegetative-reproductive cycle annually. Perennial cycles tend to be related to seasonal cycles of warm-cold (temperature climates) or wet dry periods (tropical climates). Both herbaceous and woody plants can be perennial:
- Herbaceous perennials produce shoots that grow during one season and die back during the winter or periods of drought. It may take herbaceous perennials several growing cycles before they become reproductive, and they may not flower every year, depending on the plant’s accumulation of resources during the growing cycle.
Plants survive during adverse conditions as specialized underground structures with roots and crown that remain perennial. Geophytes (bulbs, corms, rhizomes, tubers; see Chapters 15) are included in this group.
b.Woody perennials develop permanent above ground woody stems that continue to increase annually from apical and lateral buds with characteristic growth and dormancy periods. Woody perennials are trees and shrubs.
Life Cycles of Seedling Cultivars
In propagation, an individual plant that develops from a seed is referred to as a seedling whether it is an annual, biennial, herbaceous perennial, or woody perennial.
Phase I Embryonic This phase begins with the formation of a zygote. This cell grows into an embryo, which receives nourishment from the mother plant through physiological stages of development. At first, growth involves cell division of the entire embryo as it increases in size. Later, growth potential develops with a polar orientation as the embryo develops its characteristic structure. These embryonic changes are described in detail in Chapter 4.
Phase II Juvenile Seed germination initiates a dramatic change from the embryonic pattern to the developmental pattern of the young seedling. Vegetative growth is now polar, extending in two directions via the shoot and root axis. Cell division is concentrated in the root tips, shoot tips, and axillary growing points. Subsequently, the extension of the root and shoot is accompanied by an increase in volume. New nodes are continually laid down as leaves and axillary growing points are produced. Lateral growing points produce only shoots that are not competent to flower. The juvenile period is the growth stage where plants cannot flower even though the inductive flowering signals are present in the environment.
Phase III Transition The vegetative period at the end of the juvenile phase and prior to the reproductive stage is marked by subtle changes in growth and morphology. Growth tends to decrease as the plants enter the reproductive period when flowering occurs. The important point is that the developmental potential of the growing points is sensitive to particular signals, partly internal, although often dictated by cues from the environment such as changes in day length and chilling.
Phase IV Adult (or Mature) During this phase, shoot meristems have the potential to develop flower buds, and the plant produces flowers, fruits, and seeds. The duration and expression of these phases represent fundamental variation in plant development, which is analogous to comparable phases in animal development. The most conspicuous expression of phases occurs in long-lived perennial plants, such as trees and shrubs, where conspicuous differences in juvenile and mature traits may be observed in the same plant. Nevertheless, phase changes have been identified in annual plants, such as maize (61), and must be recognized as a fundamental aspect of all plant development.
The following characteristics of plant development are associated with phase change:
- Time of flowering (52, 79, 85). The age when flowering begins is the most characteristic aspect of phase change. Time of first flowering varies from days to a few months in some annuals to as much as 50 years in some perennials (Table 2–1). Usually, flowering begins in the upper and peripheral parts of the tree where shoots and branches have attained the prerequisite phase.
- Morphological expression of leaves and other structures. Leaf form in the juvenile phase sometimes differs radically from that of the adult phase (Fig. 2–2). English ivy is a classic example of phase change, as illustrated in Figures 2–3 and 2–4. Juvenile parts of apple, pear, and citrus seedlings may be very thorny, although the trait disappears in the adult phase (33, 80). Potential for regeneration (34, 80). Each phase tends to have a differing potential for regeneration. For instance, cuttings taken from the juvenile phase usually have a higher potential for rooting than do cuttings from the adult phase. An in-depth discussion of the impact of phase change on propagation is found in Chapter 16.
Organisms are named in a hierarchical system described as their taxonomy. A sample hierarchy is provided for apple (Table 2–2). The basic system for naming plants was introduced by Linnaeus (Fig. 2–4) as the Latin system of binomial nomenclature using a genus and species name for each plant (each of which are italicized). The genus describes a group of plants that are similar in morphological, biochemical, and genetic properties. The species is used to designate a population of plants within a genus that can be recognized and reproduced as a unit (51). The rules for naming plants are maintained by the International Association of Plant Taxonomists under the longstanding International Code of Botanical Nomenclature (http://ibot.sav.sk/icbn/main). In nature, individuals within one species normally interbreed freely but do not interbreed well with members of another species. Geographical isolation or some physiological, morphological, or genetic barrier prevents gene exchange between them. A true species can usually be propagated and maintained by seed but may require some control during propagation.
Cultivated plants may also be designated by binomial name even though they may be a complex hybrid rather than a distinct “natural” species (51, 72). For example, peach cultivars are variations within a recognized species Prunus persica L., but the European prune (Prunus domestica L.) is a complex hybrid that apparently developed in cultivation. Cultivars may also be derived from repeated vegetative propagation of an initial desirable mutation. The rules for naming cultivated plants are spelled out in the International Code of Nomenclature for Cultivated Plants (9).
Eastern redbud illustrates the various subgroups occurring in selected or natural populations within a species. In some cases, breeders have been able to make genetic crosses between different species or even between genera. Interspecific hybrids within a genus are designated with an “x” between the genus and species (i.e., Viburnum xburkwoodii, which is a hybrid between V. carlesii andV. utile). Intergeneric hybrids are formed between genera within a family and are designated with an “x” before the new genus name, which is a contraction of the two genera names (i.e., xFatshedera lizei is an intergeneric hybrid between Fatsia japonica and Hedera helix).
Names for new plants should be registered with the proper registration authority. The International Society for Horticultural Sciences provides a home for the Commission for Nomenclature and Cultivar Registration (http://www.ishs.org/sci/icra.htm). They provide a link to individuals or organizations that maintain the registry for a single genus or group of plants. For example, the registry for English ivy (Hedera) is maintained by the American Ivy Society, while woody plants without specific registries are handled by the American Public Gardens Association.
LEGAL PROTECTION OF CULTIVARS
In modern agricultural and horticultural industries, individual cultivars and breeding materials have commercial value and, according to law, are entitled to legal protection as is any invention made by humans (17, 40, 42, 59). The right to propagate specific cultivars that are developed through controlled selection and/or breeding programs can be protected by a number of legal devices. These allow the originator to control their distribution and receive monetary awards for their efforts.
Legal protection has been available in the United States with the passage of the Townsend-Purnell Act in 1930, which added vegetatively propagated plants to the general patenting law for inventions. Protection was provided to seed-propagated cultivars by the 1970 Plant Variety Protection Act, revised in 1994 (4). Many countries of the world have legal systems that grant protection to patents and breeders’ rights, and a large network of such programs have developed. Guidelines have been produced by the International Union for the Protection of New Varieties of Plants (http://www.upov.int/index_en.html) in 1961, 1972, 1978, and 1991 (77) and the Food and Agriculture Organization of the United Nations (38). Propagators need to be aware of the rights and obligations under these particular conditions (see Box 2.1, page 22).
GENETIC BASIS FOR PLANT PROPAGATION
The life cycle of plants begins with a single cell known as a zygote. This cell is the result of the union of male and female gametes. From this initial cell, additional cells multiply and develop the body of the plant. Living plant cells contain a nucleus embedded within the cytoplasm, all enclosed within a cell wall (Fig. 2–7). The nucleus contains the genetic material that directs growth and development by determining when particular RNAs (ribonucleic acid) and proteins are made by a cell. Chromosomes within the nucleus contain DNA (deoxyribonucleic acid) that forms the genetic blueprint for heredity. DNA is present in two other structures of the cell—chloroplasts and mitochondria. Individual characteristics and traits are associated with sequences of DNA nucleotides coded on the chromosome as genes. Genetic information is passed along from cell to cell during cell division.
PLANT HORMONES AND PLANT DEVELOPMENT
Plant hormones (phytohormones) are naturally occurring organic chemicals of relatively low molecular weight, active in small concentrations. The classic definition of a hormone is that they are synthesized at a given site and translocated to their site of action; however, there are some exceptions for plant hormones. They are specific molecules involved in the induction and regulation of growth and development. The five major plant hormones are auxin, cytokinin, gibberellin, abscisic acid, and ethylene. Additional compounds considered hormones include brassinosteroids, jasmonates, salicylic acid, polyamines, and peptide hormones. Plant hormones have great importance in propagation because they not only are part of the internal mechanism that regulates plant function, but they also can induce specific responses such as root initiation in cuttings and dormancy release in seeds. In addition to these substances, certain chemicals— some natural, others synthetic—show hormonal effects to plants. Both natural and synthetic types are classed together as plant growth regulators (PGRs).
Here is a usual set of events that occurs during hormone-induced growth and development:
- Biosynthesis of the hormone
- Transport or distribution to its site of action
- Perception of the hormone signal by its cellular receptor
- Signal transduction leading to downstream events often at the molecular (gene expression) level.
It has become evident that many types of growth and development are not controlled by a single hormone; rather there is considerable interaction and “cross-talk” often between several hormones. Often there is one principle hormone controlling development with other hormones modifying its action. For example, as discussed in Chapter 7, abscisic acid’s control over seed dormancy is modulated by gibberellin, cytokinin, ethylene, and brassinosteroid. Some of the plant hormones are present in active and conjugated forms. Conjugation is the addition of a sugar or amino acid to the chemical structure of the hormone. Conjugation may inactivate the hormone permanently, or enzymes can interconvert the hormone between conjugated and free forms through a process called homeostatic control.
Auxin was the first plant hormone discovered by plant scientists. Phototropism, where uni-directional light altered the growth of plant coleoptiles, in grass seedlings was one of the first biological systems studied by botanists including Charles Darwin (22). Fritz Went, Kenneth Thimann (82), and a number of other researchers showed that these effects could be induced by plant extracts, which were subsequently shown to contain the chemical indole-3-acetic acid (IAA). There are two biosynthetic pathways for IAA in plants (5). Primary auxin biosynthesis is via the amino acid L-tryptophan, but IAA can also be synthesized by a tryptophan-independent pathway. Most of the IAA in plant tissue is in the conjugated form using both amino acids and sugars for conjugation. Free, active IAA comprises approximately 1 percent of the total auxin content, with the remaining portion in the conjugated form. Primary sites of auxin biosynthesis include root and shoot meristems, young leaf primordia, vascular tissue, and reproductive organs including developing seeds (Fig. 2–25).
Auxin movement from cell to cell requires efflux carriers located on the plant membrane (Fig. 2–26) (83). They control polar auxin movement from plant tips (distal ends) to their base (proximal end). Cellular auxin movement and the subsequent polar gradient established between cells is important for normal development of the plant embryo as well as the shoot apical meristem.
Auxin has a major role for controlling phototrophism, inhibition of lateral buds by terminal buds (apical dominance), formation of abscission layer on leaves and fruit, activation of cambial growth, and adventitious root initiation. Auxin is the most widely used hormone in plant propagation because of its impact on adventitious rooting in cuttings (see Chapter 10) and its control of morphogenesis during micropropagation (see Chapters 17 and 18). Synthetic auxins are less susceptible to IAA-oxidase degradation and are, therefore, used more often for commercial applications. The most useful synthetic auxins, discovered about 1935, are indole-3-butyric acid (IBA) and 1-naphthalene acetic acid (NAA). IBA has been subsequently found to occur naturally, but in less abundance compared to IAA. IBA must be converted by plant tissue into IAA to function. The herbicide, 2,4-D (2,4-dichlorophenoxyacetic acid) has auxin activity and is an important inducer of somatic embryogenesis in tissue culture (see Chapter 17). Various synthetic IBA conjugates (such as its aryl ester PITB— Fig. 2–25) have been developed with good auxin activity but are not widely available or used (35). Auxins are not readily dissolved in water and must be dissolved in a solvent (ethanol, DMSO) or a base (1N NaOH) before being quickly added to water. Potassium salts of IBA and NAA (K-IBA, K-NAA) are auxin formulations that easily dissolve in water and are available commercially.
Cytokinins were discovered by Miller and Skoog at the University of Wisconsin in efforts to develop methods for growing plant cells in tissue culture (68). Through the 1940s and 1950s, researchers were frustrated because isolated plant cells and tissues grew poorly or not at all in tissue culture. At that time, tissue culture media supplemented with coconut milk (liquid endosperm) had the most stimulating effect on cell division compared to other compounds evaluated. Then Miller and Skoog inadvertently discovered that an extract from autoclaved fish sperm DNA yielded a compound that greatly stimulated cell division. This synthetic compound was called kinetin and the hormone class was called cytokinins because of their ability to stimulate cell division. Subsequently the naturally occurring cytokinins zeatin (isolated from corn endosperm) and isopentenyladenine (2iP) were found in seeds and other plant parts. These previously mentioned cytokinins along with the naturally occurring dihydrozeatin and the synthetic benzyladenine (BA or BAP) represent the aminopurine type cytokinins (Fig. 2–27). Another class of compounds—the dipheylureas—displays potent cytokinin activity but are structurally dissimilar to natural occurring cytokinins, including thiourea, diphenylurea, thidizuron (TDZ), and N-(2–chloro-4-pyridyl)n–phenylurea (CPPU). The major route for cytokinin biosynthesis is via the isoprenoid pathway with isopenteyltransferase (ipt) being the key regulatory enzyme (48). The root tip is a primary source of cytokinin, but biosynthesis also occurs in seeds (embryos) and developing leaves. In addition to free forms of cytokinin, conjugated derivatives include ribosides, ribotides, aminoacids and sugars—many of which freely interconvert. The major enzyme for cytokinin destruction is cytokinin-oxidase. Cytokinins are thought to play a regulatory role in cell division, shoot initiation and development, senescence, photomorphogenesis, and apical dominance. Cytokinins play a key role in regulating various aspects of the cell cycle and mitosis. Transgenic plants over-expressing the ipt gene show elevated cytokinin levels, reduced height, increased lateral branching, and reduced chlorophyll destruction leading to a deep green color. Tissue infected with Agrobacterium tumifaciens grow and proliferate in tissue culture independent of growth regulator application. This is because it induces elevated cytokinin levels by inserting an ipt gene from its plasmid into the plant’s genome. The interaction of auxin and cytokinin is one of the primary hormonal relationships in plant growth and development as well as plant propagation.
A high auxin:cytokinin ratio favors rooting, a high cytokinin:auxin ratio favors shoot formation, and a hhig level of both favors callus development (see Chapter 9 on rooting and Chapter 17 on micropropagation).
Gibberellins (69) were discovered before World War II by Japanese scientists trying to explain the abnormally tall growth and reduced yield of rice infected by the fungi Gibberella fukikuori (perfect stage) or Fusarium moniliformne (imperfect stage). An active ingredient was extracted from the fungus and its chemical structure was determined as gibberellins (named after the fungus). Subsequently, gibberellins were found to be naturally occurring hormones in plants. All gibberellins are cyclic diterpenoids and named for their structure not their activity. More than 100 forms of gibberellins have been found in plants but only a few are physiologically active. The most important naturally occurring active gibberellins include GA1, GA4, GA7 (Fig. 2–28). Depending on the plant, they will tend to make either GA1 or GA4 as their primary gibberellin. Gibberellic acid (GA3) is the gibberellin found in fungi and is the most important commercial product. Biosynthesis of gibberellins (73, 76) starts with mevalonate (an important precursor for many secondary compounds in plants) and proceeds via the iosprenoid pathway. Its biosynthesis is a coordinated process involving the plastids, endoplasmic reticulum, and cytosol. Numerous enzymes are regulated during gibberellin biosynthesis, but GA20 oxidase appears to especially important. Active gibberellins are inactivated by GA2 oxidase. Gibberellins can also be sugar conjugated as previous discussed with other hormones. Gibberellins are made in developing seeds and fruits, elongating shoots, and roots. Gibberellins are the primary hormone controlling plant height. Gibberellin mutants impaired for gibberellin biosynthesis are dwarfed compared to wild type plants, demonstrating the importance of gibberellins for shoot elongation. Several commercially available gibberellin biosynthesis inhibitors, including ancymidol, cycocel, paclobutrazol (Bonzi), and uniconizole (Sumagic), are important plant growth regulators used to control plant height during greenhouse pot and bedding plant production. Gibberellins also play a role in plant maturation and in triggering flowering. Gibberellins are particularly important during seed germination, where the antagonistic interactions between gibberellin and abscisic acid are involved in dormancy release and germination.