Tissue Culture Of Trees

Woody plants provide many challenges to the tissue culturist. Although there are many excellent tissue culture books and manuals available, these are generally strongly biased towards herbacious crops. Consequently, they often do not pay sufficient attention to the problems that specifically apply to in vitro culture of tree species. Culture of the latter often poses problems which are either absent or of lesser significance when culturing herbacious species. When trees in the field are used as explant source, the problems can be especially severe. For example, the physiological condition of the explants is difficult to control because of variation in weather and biotic factors. Furthermore, it is often difficult to obtain explants free of contaminants from field grown trees. Lack of genetic uniformity and maturation are additional problems one often has to deal with when culturing tree cells or tissues. These problems are emphasized in this text. In vitro culture of trees is not viewed in isolation. It is considered in conjunction with breeding, traditional cloning and other common tree improvement techniques. The text discusses theoretical as well as practical aspects of the in vitro culture of trees.

This symposium is the third in a series featuring the propaga tion of higher plants through tissue culture. The first of these symposia, entitled "A Bridge Between Research and Application," was held at the University in 1978 and was published by the Technical Information Center, Department of Energy. The second symposium, on "Emerging Technologies and Strategies," was held in 1980 and pub lished as a special issue of Environmental and Experimental Botany. One of the aims of these symposia was to examine the current state of-the-art in tissue culture technology and to relate this state of technology to practical, applied, and commercial interests. Thus, the third of this series on development and variation focused on embryogenesis in culture: how to recognize it, factors which affect embryogenesis, use of embryogenic systems, etc.; and variability from culture. A special session on woody species again emphasized somatic embryogenesis as a means of rapid propagation. This volume emphasizes tissue culture of forest trees. All of these areas, we feel, are breakthrough areas in which significant progress is expected in the next few years.

1 John H. Dodds The culture offragmen ts of plant tissue is not a particularly new science, in fact as long ago as 1893 Rechinger (1893) described the formation of callus on isolated fragments of stems and roots. The culture of plant tissues in vitro on a nutrient medium was performed by Haberlandt (1902), however, his attempts were unsuccessful because he chose too simple a medium that lacked critical growth factors. Over the last fifty years there has been a surge of development in plant tissue culture techniques and a host of techniques are now avail able (Dodds and Roberts, 1982). The major areas are as follows. Callus Culture Callus is a rather ill-dermed material. but is usually described as an un organised proliferating mass of tissue. Although callus cultures have a great deal of potential in the biotechnological aspects of tissue culture, i.e. secondary product formation, they are not very suitable for plant propagation. The key reason for their unsuitability is that genetic aber rations occur during mitotic divisions in callus growth (D'amato.l965). The aberrations can be of a major type, such as aneuploidy or endo reduplication. It follows therefore that the genetic status of the re generated plants is different from that of the parent type. In general terms this genetic instability is undesirable, but there are occasions when a callus stage can be purposely included to diversify the genetic base of the crop.

2. 2. Plant materials 2. 3. Pregrowth conditions 2. 4. Cryoprotectant treatment 2. 5. Freezing 2. 5. 1. Slow freezing 2. 5. 2. Rapid freezing 2. 5. 3. Droplet freezing 2. 6. Storage 2. 7. Thawing 2. 8. Viability testing 2. 9. Post-thaw regrowth 3. EXAMPLES OF CRYOPRESERVATION OF WOODY PLANT MATERIAL 4. POTENTIAL APPLICATION OF CRYOPRESERVATION IN TREE IMPROVEMENT 17. NURSERY HANDLING OF PROPAGULES - J. A. Driver, and 320 G. R. L. Suttle 1. INTRODUCTION 2. COMMERCIAL NURSERY NEEDS VS. LABORATORY PRACTICE 3. SEASONALITY OF GROWTH AND PRODUCTION CYCLES 4. MICROPROPAGATION OPTIONS 4. 1. Trends in commercial micropropagation 4. 1. 1. Contract micropropagation 5. FACTORS AFFECTING SURVIVAL AND GROWTH 5. 1. Hardening of propagules in vitro 5. 2. Greenhouse considerationS------ 5. 3. Field planting 5. 4. New approaches: Direct field rooting 5. 4. 1. Pretreatment in vitro 5. 4. 2. Root induction 5. 4. 3. Field placement 18. MYCORRHIZAE - R. K. Dixon, and D. H. Marx 336 1. INTRODUCTION 2. ROLE OF MYCORRHIZAE IN TREE GROWTH AND DEVELOPMENT 3. PRODUCTION AND APPLICATION OF ECTOMYCORRHIZAL FUNGUS INOCULUM 3. 1. Bareroot stock 3. 2. Container-grown stock 4. FIELD TRIALS WITH ECTOMYCORRHIZAL PLANTING STOCK 5. PRODUCTION AND APPLICATION OF ENDOMYCORRHIZAL INOCULUM 6. FIELD TRIALS WITH ENDOMYCORRHIZAL 7. RESEARCH OPPORTUNITIES 8. SUMMARY 351 19. TISSUE CULTURE APPLICATIUN TO FOREST PATHOLOGY AND PEST CONTROL - A. M. Diner, and D. F. Karnosky 1. INTRODUCTION 2. HOST AND PATHOGEN: CULTURE AND CHALLENGE 2. 1.

Micropropagation has become a reliable and routine approach for large-scale rapid plant multiplication, which is based on plant cell, tissue and organ culture on well defined tissue culture media under aseptic conditions. A lot of research efforts are being made to develop and refine micropropagation methods and culture media for large-scale plant multiplication of several number of plant species. However, many forest and fruit tree species still remain recalcitrant to in vitro culture and require highly specific culture conditions for plant growth and development. The recent challenges on plant cell cycle regulation and the presented potential molecular mechanisms of recalcitrance are providing excellent background for understanding on totipotency and what is more development of micropropagation protocols. For large-scale in vitro plant production the important attributes are the quality, cost effectiveness, maintenance of genetic fidelity, and long-term storage. The need for appropriate in vitro plant regeneration methods for woody plants, including both forest and fruit trees, is still overwhelming in order to overcome problems facing micropropagation such as somaclonal variation, recalcitrant rooting, hyperhydricity, polyphenols, loss of material during hardening and quality of plant material. Moreover, micropropagation may be utilized, in basic research, in production of virus-free planting material, cryopreservation of endangered and elite woody species, applications in tree breeding and reforestation.

2. IMPORTANCE OF NITROGEN METABOLISM 2. 1. Range of naturally occurring nitrogenous components in forest trees 2. 2. Gene expression and mapping 2. 3. Metabolic changes in organized and unorganized systems 2. 4. Nitrogen and nutrition 2. 5. Aspects of intermediary nitrogen metabolism 3. NITROGEN METABOLISM IN GROWTH AND DEVELOPMENT 3. 1. Precultural factors 3. 2. Callus formation 3. 3. Cell suspensions 3. 3. 1. Conifers 3. 3. 2. Acer 3. 4. Morphogenesis 3. 4. 1. Nitrogen metabolism of natural embryos 3. 4. 2. Somatic embryogenesis 3. 4. 2. 1. Sweetgum (Liquidambar styraciflua) 3. 4. 2. 2. Douglar-fir and loblolly pine 3. 4. 3. Organogenesis 4. OUTLOOK 11. CARBOHYDRATE UTILIZATION AND METABOLISM - T. A. Thorpe 325 1. INTRODUCTION 2. NUTRITIONAL ASPECTS 3. CARBOHYDRATE UPTAKE 4. CARBOHYDRATE METABOLISM 4. 1. Sucrose degradation 4. 2. Metabolism of other carbon sources 4. 3. Hexose mobilization and metabolism 4. 3. 1. Cell cycle studies 4. 3. 2. Growth studies 4. 3. 3. Organized development 4. 4. Cell wall biogenesis 4. 4. 1. Primary cell walls 4. 4. 2. Cell wall turnover 4. 4. 3. Secondary cell walls 4. 5. Carbon skeleton utilization 5. OSMOTIC ROLE 6. CONCLUDING THOUGHTS 369 12. THE USE OF IN VITRO TECHNIQUES FOR GENETIC MODIFICATIO~FOREST TREES - E. G. Kirby 1. INTRODUCTION 2. IN VITRO SELECTION 2. 1. Natural variation 2. 2. Induction of variation 2. 3. Selection techniques 2. 4. Plant regeneration 2 . • 5. Applications x 3. SOMATIC HYBRIDIZATION 3. 1.