World Journal of Microbiology and Biotechnology, 8, 369-377

Review

Progress in the biotechnology of trees N. Hammatt An increasing world population and rise in demand for tree products, especially wood, has increased the need to produce more timber through planting more forest with improved quality stock. Superior trees are likely to arise from several sources. Firstly, forest trees can be selected from wild populations and cloned using macropropagation techniques already being investigated for fruit tree rootstocks. Alternatively, propagation might be brought about in vitro through micropropagation or sustained somatic embryogenesis, with encapsulation of the somatic embryos to form artificial seeds. Tree quality could be improved through increased plant breeding and it is likely that experienced gained, to date, in the breeding of fruit species will be useful in devising strategies for forest trees. Since the development of techniques to regenerate woody plants from explant tissues, cells and protoplasts, it is now feasible to test the use of tissue culture methods to bring about improvements in tree quality. Success has already been achieved for tree species in the generation of somaclonal and protoclonal variation, the formation of haploids, triploids and polyploids, somatic hybrids and cyhrids and the introduction of foreign DNA through transformation. This review summarizes the advances made so far in tree biotechnology, and suggests some of the directions that it might take in the future.

Key words: Clonal propagation, genetic improvement, plant breeding, plant regeneration.

We depend on trees as sources of construction materials, fuel and paper, fruits, oils, and medicines. Trees are also used ornamentally. Forests play an important role in maintaining the global hydrological cycle and in controlling atmospheric CO2, an important greenhouse gas. A third of the Earth's land mass is covered by forest, while a further small area is covered by commercial fruit orchards. Broadleaved tropical rain forests comprise a third of this forest resource, and coniferous boreal forests in the northern hemisphere account for a further 20%. A fifth consists of open forests, with scrub, declining forest and planted agricultural land making up the final 20% (Forestry Industry Committee of Great Britain, 1987). While demand for hardwood timber for construction and pulping may lead to a loss of up to 3% of tropical rainforests per annum, a much greater area of tree cover in the Third World is removed annually for fuel and agriculture. Further rapid increase in global population will continue to place immense pressure on the world's supplies of trees. Thus there is an immediate need for research to increase the productivity of trees. Fundamental to this objective will be

N. Hammatt is with Horticulture Research International, East Mailing, West Mailing, Kent, ME19 6B J, UK.

the development of procedures to clone trees with good field potential. Such superior stock is likely to derive not only from specimens collected from wild populations, but also from the genetic improvement of existing trees. The aim of this review is to summarize progress to date in techniques for the propagation and genetic improvement of trees based on tissue culture and conventional breeding. Owing to the large amount of information on some of the subjects considered, this article will provide representative examples of published papers in a given field, from which the reader should be able to extract further information through the references cited.

Tree Breeding Unlike timber species, fruit trees have been significantly and progressively improved through conventional plant breeding. Not only has attention been paid to the flavour, colour and keeping qualities of fruit, but the reliability of cropping has also become of ever increasing importance in commercial orchards (Alston & Tobutt 1989). Precocity in cropping, resulting from a reduction in the vegetative phase of a variety or juvenile phase of a selection, has been an especially important breeding objective. Not only can the

9 1992 Rapid Communications of Oxford Ltd bVorld Journal of Mwroblology and Bio~echnology, Vol 8, 1992

3~}

N. Hammatt

genotype of the scion affect precocity, but budding onto dwarfing rootstocks such as M.9 or M.27, in the case of apple, can reduce the time to fruiting (Tydeman & Alston 1965; Quinlan & Tobutt 1990). Selection of dwarf or semi-dwarf types has significantly improved cropping, and a number of genes that influence growth habit have been identified already in apples and pears (Lapins 1969, Lapins & Watkins 1973; Alston 1976; Tobutt 1985). Plant breeding techniques have a major role to play in the improvement of forest trees, and it is likely that experience with fruit trees will be of value in achieving this objective. Despite considerable research, most forest planting stock still derives from genetically undefined seed origins, with the possible exception of poplars (Populus spp.), Eucalyptus and willows (Salix spp.). This has arisen chiefly because of the long juvenile period and the length of time required to investigate fully the performance of promising selections derived from crosses. Breeding could become more rapid if, as has been achieved for bamboos (Nagauda et al. 1990; Chambers et al. 1991), micropropagation can be used to speed up the onset of flowering in trees. Early selection of superior seedlings could be facilitated if markers can be found in juvenile trees that predict aspects of the adult phenotype. In apple, for instance, the percentage of rootbark and the number of stomata can be used to predict the likely effects that apple seedlings would have on scion dwarfing, if they were subsequently used as rootstocks (Beakbane & Thompson 1939; Beakbane & Majumber 1975). Similarly, the isoenzyme GOT.1 (glutamate oxaloacetate transferase) can be used as an indicator of S incompatibility alleles in apple seedlings (Manganaris & Alston 1987). A key objective in forest breeding will be to enhance timber production. Increased vigour, a quantitative trait, might be brought about by exploitation of novel germplasm, as in rootstocks and scions of apple, in which a range from very vigorous to dwarfing types are available. Alternatively, vigour might derive from heterosis or from the elevation of ploidy levels as seen in many other crops. Improved resistance to insect browsing and pathogens should also result in increased productivity. The diversion of primary assimilates into wood could also result from the genetic manipulation of growth habit, in particular, reduction in branching, thought to be a more qualitative trait, which has already been achieved in top fruits such as apple (Quinlan & Tobutt 1990). It may be possible to plant such trees at a greater density and thereby increase timber yield per unit area. Breeding less spreading trees may also have benefits to species such as wild cherry (Prunus avium), in which close proximity of adjacent crowns can lead to the decline of a stand. The production of sterile trees should result in enhanced vegetative growth as a consequence of the reduced energy expenditure involved in fruit development. Initially, this objective might be achieved through the formation of

370

World ]ournal of Microbiology and Bio~echnology, Vol 8, 1992

triploid trees produced by crossing diploids with tetraploids. Triploids, which are usually sterile, do not normally produce fruit and are more likely, therefore, to divert primary assimilates into wood, a desirable trait for forest trees. When fruits are produced, as with Citrus, Musa and Malus, they are often larger than their diploid counterparts, and usually devoid of seeds. In future breeding programmes, the quality of wood also needs to be considered. For instance, veneer quality timber, which attracts the highest premium, usually derives from slower growing trees, while the length and density of the fibres and the lignin content of wood influence its pulping qualities.

Genetic Manipulation In Vitro Plant Regeneration

The advent of techniques of plant regeneration has permitted the recovery of novel plants from genetically manipulated cells, tissues and protoplasts in an everincreasing range of herbaceous plants. Since the attention of tissue culturists has extended to include trees only very recently, less is known about how to regenerate plants from somatic tissues of woody plants. Consequently, techniques for genetic manipulation in vitro have resulted in only a few genetically novel trees. Plantlets can be recovered from explant material, either via somatic embryogenesis or through the formation of adventitious shoots, as has been achieved with organ and tissue explants from several timber and fruit tree species (James 1987; Jones 1992a; Hassig 1989; Valobra & James 1990). Plants have also been regenerated from protoplasts of Citrus (Vardi et al. 1982; Sim et al. 1988), and several temperate Rosaceous top fruit species including pear (Ochatt & Power I988a), apple (Patat-Ochatt et al. 1988) and both sour cherry (Ochatt & Power 1988b) and 'Colt' cherry (Prunus avium x pseudocerasus; Ochatt et al. 1987). In forest trees, plants have been obtained from protoplasts of broadleaved species such as elm (Sticklen et aL 1986) and poplar (Russell & McCown 1988) and conifers such as Picea glauca (Attree et al. 1989) and Larix x eurolepis (Klimaszewska 1989). It is clear that success in obtaining adventitious shoots depends primarily on the genotype used and thus, until this problem of genotype dependence can be overcome, genetic manipulations in vitro will be restricted to responsive genotypes. However, conventional plant breeding techniques may be useful in facilitating gene flow from recombinants produced in vitro into recalcitrant germplasm. Somaclonal Variation

The process of adventitious regeneration often results in heritable mutations referred to as somaclonal variation, only

Biotechnology of trees a small proportion of which is useful. In trees, Lester & Berbee (1977) first detected somadonal variation in height, branching and leaf traits amongst regenerant poplar plants, while Ostry & Skilling (1988) later described poplar somaclones more resistant to the leaf spot fungus Septoria musiva. Similarly in P. deltoides (eastern cottonwood), both enhanced and reduced resistance to leaf rust, caused by Melampsora medusae, was detected in regenerated plants (Prakash & Thielges 1989). In fruit trees Hammerschlag (1988) produced peach plantlets which were more resistant than the parental tree to the bacterial pathogen Xanthomonas campestris var. pruni, while Donovan (1991) generated apple trees resistant to fireblight caused by Erwinia amylovora in the susceptible scion variety 'Greensleeves'. Salt-resistant somaclones have been obtained from explants and protoplasts of the cherry rootstock 'Colt' (Ochatt & Power 1989) and protoplasts of Citrus (Ben-Hayyim & Goffer 1989). In most cases the genetic basis and long-term potential for useful somaclonal variants have not yet been adequately assessed.

Manipulation of Ploidy Endosperm, derived from double fertilisation in Angiosperms, is triploid, and therefore a possible, alternative source to breeding of triploid plants. Direct regeneration of triploid plants from endosperm has already been achieved for several trees including Citrus (Wang & Chang 1978), sandalwood (Santalum album; Lakshmi Sita et al. 1980) and walnut (]uglans regia; Tulecke et al. 1988). It is possible that tetraploids will be more vigorous than their diploid parents, and they could be used as parents in crosses with diploids to generate triploids. While conventional treatments with colchicine have resulted in polyploid production in species such as apricot (Prunus armeniaca; Lapins 1975), it has also been possible to increase ploidy in vitro. Thus James et al. (1987) cultured micropropagated shoots of the triploid cherry rootstock 'Colt' in a rooting medium containing auxin and colchicine and produced some hexaploid roots, which gave rise to hexaploid shoots. The latter may be more effective as rootstocks than their triploid counterparts. Reduction to the haploid state also has uses to breeders and geneticists (Radojevi4 & Kovoor 1985). In some species, including apple, it has been possible to generate haploids by sexual crossing (Lespinasse et al. 1983). The recovery of haploids through regeneration from germ cells of trees is hampered by the very short time period, annually, during which plants are flowering and producing suitable explant material. However, haploids have been produced by this means, for instance, in apple (Fei & Xue 1981; Wu 1982), horsechestnut (Aesculus hippocastanum; Radojevi4 1978) and red horsechestnut (A. carnea; Radojevi4 et al. 1989).

Genetic Transformation The rapid advances in the practice and application of plant transformation have been reviewed extensively (e.g. Gasser & Fraley 1989). As with other areas of biotechnology, the routine generation of transgenic trees will not become a reality until procedures for plant regeneration are better understood. It is encouraging, however, to find that many tree species are susceptible to infection by Agrobacterium, and that foreign DNA can be incorporated into tree genomes and expressed. In some woody species, for which regeneration techniques have been developed, transgenic plants have been recovered from transformed tissues. Both Fillatti et al. (1987) and de Block (1990) have produced transgenic poplar trees more tolerant of herbicides, a trait which would facilitate the chemical control of weeds in the early management of ctonal plantations. Transgenic plants have also been obtained in walnut (Juglans regia; McGranahan et al. 1988), apple (James et al. 1989), Allocasuarina (Phelep et al. 1991), plum (Mante et al. 1991) and peach (Smigocki & Hammerschlag 199I). The preparation of protoplasts results in the exposure of the plasmalemma to external agents. Following electroporation or treatment with polyethylene glycol (PEG), or both of these treatments applied together, protoplasts introduced into a suspension of naked DNA can take up the foreign material, and expression of easily scored genes has been observed. Transient expression of the bacterial GUS gene (fl-glucuronidase), has already been achieved in protoplasts of white spruce (Picea glauca; Wilson et al. 1989) and alder (Alnus incana; S~guin & Lalonde 1988), while the enzyme CAT (chloramphenicol acetyl transferase) has been transiently expressed in protoplasts of P. glauca (Bekkaoui et al. 1988; Wilson et al. 1989). Expression of the luciferase gene, luc, has been detected in protoplasts of Douglas fir (Pseudotsuga menziesii) and loblolly pine (Pinus taeda) (Gupta et al. 1988). Recently, transgenic plants of Citrus have been obtained from transformed protoplasts (Vardi et al. I990). Following further progress in the development of regeneration techniques for woody plants and in the understanding of factors that affect virulence of Agrobacteria to trees, examples of successful genetic transformation of trees are likely to become more common. Genes that confer resistance to insects, such as those that code for cowpea trypsin inhibitor or the crystal toxin proteins of Bacillus thuringiensis, if transferred to trees, could provide an important contribution to the management of orchards and forest plantations. Furthermore, expression of viral coat proteins could assist in preventing yield reductions caused by systemic viruses. A common criticism of the use of such genes in isolation, however, is that the resulting qualitative resistance could be overcome rapidly by the pest. Furthermore, it is possible that resistance to browsing insects, the primary consumers, will disturb the food chain

World Journal of Microbiology and Biotechnology, Vol 8, 1992

3 71

N. Hammatt and thus serve to destabilize the woodland ecosystem, and reduce its overall productivity. For these reasons, combined with public concern over the release of genetically manipulated organisms and the longevity of the crop, it is questionable whether single gene resistance, introduced in this way, will be of long-term value in trees. Quantitative resistance to pests is probably silviculturally and environmentally more acceptable. Traditionally, quantitative traits have been introduced into a crop through techniques of plant breeding. Thus, as with other plants, if transformation is to be useful for the generation of pest resistant trees, it will be necessary to transfer a larger number of genes than has so far been achieved. In the meantime, transformation techniques could provide a useful means to transfer more qualitative features, such as branching habit and herbicide resistance, the latter facilitating the establishment of clonal plantations. It is likely that transformation will provide a valuable tool in the utilization of molecular approaches, such as the use of antisense sequences and overexpression of genes, to investigate the genetic control of juvenility, which has important implications for growth rates and ease of rooting of conventional cuttings, and to reduce lignin production which would improve the pulping qualities of timber.

Somatic Hybridization and Cybrid Production The fusion of protoplasts and recovery of somatic hybrid and cybrid plants from the resulting fusion products have been documented in many review articles (e.g. Bravo & Evans 1985). The process offers unique opportunities to combine two sexually compatible or incompatible, diploid complements into novel tetrap[oids. Since both protoplasts involved in a heterofusion event have a full complement of nuclear and cytoplasmic genes, the limitations of maternal inheritance of cytoplasmic traits, can be overcome. Since it is possible to eliminate either the nuclear genome by gamma irradiation or the cytoplasmic genome by compounds such as iodoacetate, plants with unique nuclear-cytoplasmic combinations can be produced, as shown, for instance, in Brassica spp. (Barsby et al. 1987). With trees, almost all of the somatic hybrids produced to date have been between Citrus species and close relatives (Grosser et al. 1990). Clearly this success derives from the ease with which plants can be regenerated from Citrus protoplasts and this considerable advantage has also led to a report of cybrid trees within the genus Citrus, and between Citrus spp. and Poncirus trifoliata (Vardi et al. 1987). Besides Citrus, Ochatt et al. (1989) have described a somatic hybrid between wild pear (Pyrus communis) and 'Colt' cherry. The authors have suggested that the pentaploid somatic hybrid may be a useful rootstock for both pears and cherries.

372

World ]ournal of Microbiology and Biotechnology, Vol 8, 1992

Selection and Clonal Propagation of Trees Selection of Superior Individuals Given the long delay expected before genetically improved forest trees become available for planting, it will be necessary meanwhile to clone pre-existing, superior trees. An initial problem encountered involves the identification of those features of a superior adult phenotype that can be attributed to the genotype, and those which result from its environment. A possible route to circumvent this problem would be to use molecular probes to screen the genome, but such diagnostic tools are not yet available. In the meantime, the most realistic approach will be to select superior phenotypes and assume that, at worst, a tree with a superior phenotype is a genotype that responded well to good management practices; while, at best, in the absence of human interference, the genotype is totally responsible for its enhanced field performance. A further difficulty is encountered in the collection of material from large, old trees for propagation. In many species, such as oaks (Quercus spp.) and limes (Tilia spp.), epicormic shoots or root suckers provide a possible source of material to propagate. In others, however, such as ash (Fraxinus excelsior), the most accessible shoots are usually high-up and thus difficult to reach, necessitating the use of cumbersome climbing and cutting equipment. A more drastic approach might be to lop or even fell the tree and use the resulting stump sprouts. Since woody material obtained from adult trees is usually difficult or impossible to root, taking cuttings directly from the parent tree would be a wasteful procedure to produce nursery stock plants. A more effective method would be to graft buds onto suitable rootstocks as is traditionally practised in the propagation of fruit trees (Howard 1987) and for a range of ornamental trees. Thus, in establishing a collection of wild cherry (Prunus avium) at East Malling, bud grafting on to 'Colt' cherry rootstocks has been successful. While clonal rootstocks would enhance the uniformity of the propagated stock, such clones would not be available in early experiments with most species, and grafting onto a mixed seedling population would be more realistic, as achieved at East Malling with ash (F. excelsoir). Clonal Propagation Having collected and accessioned material, a method to propagate the material needs to be selected. As a tree ages and becomes mature, its growth rate usually slows and it becomes progressively more difficult to propagate from conventional cuttings. In most species, however, it is only at this stage that its potential silvicultural value becomes apparent. While propagating adult fruit tree scions by grafting onto clonally produced rootstocks is financially viable, this approach is economically the least attractive way to produce forest planting stock. It would be more profitable

Biotechnology of trees to propagate forest trees by cuttings, necessitating methods to recapture rooting ability lost when the tree aged. Many clonal top fruit rootstocks are produced by rooting cuttings obtained from regularly pruned hedges and stoolbeds or by layering (Howard I987). It has been noted that the enhanced rooting ability, prolonged leaf retention and enhanced production of short axillaries, by shoots produced by each method, are characteristic of the juvenile phase of growth. These management methods may thus result in partial rejuvenation of the tissues and could have an important role to play in the mass cloning of forest trees. An altemative approach to cloning is provided by micropropagation. This process is based upon the principle that a bud taken from a given specimen, and cultured on a nutrient medium containing growth regulators, usually at least one cytokinin, develops into a shoot, and its axillary buds grow out to form side branches. These side branches can be rooted or transferred to flesh multiplication medium in which axillary buds again grow out. This process can be carried out indefinitely to produce a limitless number of shoots. Following success with large numbers of herbaceous species, information is now slow[y being gathered on the micropropagation of an increasing range of fruit and timber trees. For forest trees, most success has been reported in conifers, such as spruces (Piceaspp.), pines (Pinus spp.; Baxter et al. 1989) and firs (Abies spp.), and colonizing broadleaved species such as birches (Betula spp.; e.g. Valobra & James 1990) and alders (A]nus spp.; e.g. Barghchi I988; P6rinet et a]. 1988). For many important hardwood genera, such as tropical Dipterocarps (Linington 1991), maples (Acer spp.; Preece eta]. I991a,b), ashes (Fraxinus spp.; Hammatt & Ridout 1992) and oaks (Quercus spp.; e.g. Manzanera & Pardos 1990), little information on micropropagation has yet been published. Furthermore, most studies have used juvenile explants from genetically variable, untested seedlings or embryos, rather than trees that have reached maturity and whose long-term performance has been assessed. Based on published data, explants from mature trees are likely to be more difficult to establish in vitro owing to excessive polyphenol oxidation and rapid necrosis of the explant, but in some species this problem has been overcome (e.g. Vieitez & Vieitez 1980; Rugini & Fontanazza 1981; Marks & Simpson 1990; Linington 1991). Systemic microbes, particularly bacteria, have also often hindered micropropagation of adult trees. Remedies for this problem have included regular screening of cultures using microbial culture media, with the subsequent removal of contaminated shoots from experiments (Webster & Jones 1989a) and culture on media with antibiotics (Cornu & Michel 1987; Wilson & Power 1989) or bacteriostatic compounds such as phloroglucinol as used with the cherry rootstock, P. avium FI2/1, (Hammatt, unpublished). Little is known about the effects of viruses upon micropropagation,

although meristem culture has long been used as a means to eliminate virus from crops, including gooseberry (Jones & Vine 1968).

Field Performance of Micropropagated Trees Few laboratories have yet reported on the field performance of micropropagated trees. Apple has probably been the most studied species. Irrespective of the origin of the rootstock, composite apple trees with micropropagated scions have been shown to be more vigorous and productive of wood than their conventional counterparts, and usually experienced a delay in cropping. (Webster et aI. I986; Jones & Hadlow 1989; Mackenzie 1989; Zimmerman & Miller I991). When the rootstock was derived by micropropagation, more suckers and burr knots were observed (Navatel et al. 1988; Jones & Hadlow 1989). The wood of micropropagated composite apple trees comprised a higher proportion of fibres and fewer vessels than conventional trees, and their mid-season vessels were wider in diameter (Mackenzie, 1989). Micropropagation therefore also might be useful in widening the range of uses of timber from forest trees. With the difficult to propagate M.9 rootstock of apple, Webster & Jones (1989a) observed a gradual increase in rooting potential in vitro at each subculture. In addition, the same authors found that cuttings taken from micropropagated trees of M.9 (Webster & Jones I989b), and also several rootstocks of pear (Pyrus communis; Jones & Webster 1989), were easier to root than cuttings from conventional adult sources. Likewise, cuttings taken from micropropagated plants of Ficus benjamina were easier to root and produced more vigorous plantlets than those taken from conventionally propagated stockplants (Kristiansen 199i). In the plum rootstock 'Pixy' (Prunus insititia), the improved rooting of cuttings from micropropagated trees was still evident 9 years after the establishment of the donor hedges (Howard et al. 1989). If enhanced growth rates and improved conventional vegetative propagation can be repeated with timber species, micropropagation could have important implications for the development of a clonal forestry industry. Apparent Rejuvenation Given that vigour, delayed flowering and enhanced rooting ability are characteristics of the juvenile phase of tree growth, it is possible that micropropagation results in the rejuvenation of mature tissues. This is supported by the observation that micropropagated plants of Sequoiadendron giganteum expressed a protein found in juvenile but not mature trees (Bon & Monteuuis 1991). Jones & Hadlow (1989) concluded that the juvenile characteristics observed with micropropagated apple probably did not result from virus elimination since their study utilized virus-free material. Furthermore, the increase with time of rooting ability of

World journal oj Microbiology and Biotechnology, Vol 8, 1992

3 73

N. Hammatt

M.9 in vitro was gradual and not sudden (Webster & Jones 1989a) as might have been expected if systemic viruses had been removed. Genetic change was another unlikely explanation, since the rnicropropagated trees appeared to be true-to-type (Jones & Hadlow 1989). The observation that apple rootstocks derived from cuttings from micropropagated stock plants lost the suckering habit typical of directly transplanted micropropagules suggests that changes observed after micropropagation with apple are physiologically and not genetically based (Jones 1992b). Automation of Micropropagation Micropropagules, particularly for forest planting, will initially need to compete with the low cost of conventional seedlings. However, since micropropagation is presently costly and labour-intensive, the technique may attract greater favour as a means to recapture ease of rooting of conventional cuttings, that can be exploited in a conventional nursery system, than as a means to produce planting stock itself. However, micropropagation still could become economically viable if significant cost reductions could be achieved through automation (Levin & Vasil 1989), which is being investigated, for instance, with shoot cultures of radiata pine (Pinus radiata; Aitken-Christie & Jones 1987). Sustained secondary somatic embryogenesis, coupled with liquid culture, as practised with herbaceous plants (e.g. Ammirato 1983), and artifical seed technology (Teasdale & Buxton 1986; Durzan 1988; Bapat & Rao 1988), may also facilitate the automation of propagation. However, in many species, plantlets recovered from somatic embryos have lost clonal fidelity owing to induced mutation during tissue culture (e.g. Barwale & Widholm 1987). While this problem may limit somatic embryogenesis as a cloning technique, mutation rates could be limited by careful attention to in vitro manipulations (Karp 1989). Most of the research on somatic embryo production in trees has used genetically variable embryos or seedlings as a source of explants [e.g. Norway spruce (Picea abies; von Arnold & Hakman 1988) and white ash (Fraxinus americana; Preece et al. 1989)]. In order to exploit this technique to clone superior trees then, unless true-to-type embryos can be obtained for instance through induced apomixis, somatic embryos will have to be induced from mature tissues. This has only been achieved to date in sessile oak, beech (J6rgensen 1988), evergreen oak (F6raud-Keller & Espagnac 1989), horsechestnut (J6rgensen 1989) and willow (Gr6nroos et al. 1989). Composition of Clonal Forests Unlike annual species, the long-lived nature of a forest will serve to increase the likelihood that the crop will be affected by pathogens or pests. Thus clona] forests will have to be planted with a range of genetically different, superior individuals. Estimates have suggested that a mixture of 20 to 40 genotypes might be satisfactory.

3 74

World Journal of Microbiology and Biotechnology, Vol 8, 1992

Conclusions It is clear that, despite considerable advances in research, plant biotechnology has yet to make a contribution to the improvement of most tree species. This has resulted, to some extent, from the time delay in the development of techniques to rejuvenate and clone mature trees, and to regenerate plants from tissues and cells, as well as the long time required for conventional breeding. In most cases, only isolated, responsive, model genotypes have been considered. Consequently, one of the challenges for plant biotechnologists working with woody species will be to understand how to micropropagate and regenerate plants from an extended range of tree genotypes. Results from experiments to date, to manipulate woody plants genetically, do not appear to deviate greatly from those obtained from herbaceous plants. Thus, plant regeneration has given rise to potentially useful somaclonal and protoclonal variation, and has resulted in the generation of beneficial changes in ploidy levels, somatic hybrids and cybrids following protoplast fusion, and the transfer of foreign DNA to trees and its expression. It is likely, therefore, that after the long time delay required to develop the techniques of plant biotechnology, and to assess the performance of the resulting plant material, such approaches will eventually have a significant impact upon the quality of planting stock for both fruit and forest trees.

References Aitken-Christie, J. & Jones, C. 1987 Towards automation: Radiata pine shoot hedges in vitro. Plant Cell, Tissue and Organ Culture 8, 185-196. Alston, F.H. 1976 Dwarfing and lethal genes in apple progenies. Euphytica 2, 505-514. Alston, F.H. & Tobutt, K.R. 1989 Breeding and selection for reliable cropping in apples and pears. In Manipulation of Fruiting, ed Wright, C.J. pp. 329-339. London: Butterworth. Ammirato, P.V. 1983 The regulation of somatic embryo development in plant cell cultures: suspension culture techniques and hormone requirements. Bio/Technology 1, 68--74. Attree, S.M., Dunstan, D.I. & Fowke, L.C. 1989 Plantlet regeneration from embryogenic protoplasts of White Spruce (Picea glauca). Bio/Technology 7, 1060--1062. Bapat, V.A. & Rao, P.S. 1988 Sandalwood plantlets from 'Synthetic seeds'. Plant Cell Reports 7, 434-436. Barghchi, M. 1988 Micropropagation of Alnus cordata (Loisel.). Plant CeK Tissue and Organ Culture 15, 233-244. Barsby, T., Chuong, P.V., Yarrow, S.A., Sau-Ching, W., Coumans, M., Kemble, R.J., Powell, A.D., Beversdorf, W.D. & Pauls, K.P. 1987 The combination of Polima cms and cytoplasmic triazine resistance in Brassica napus. Theoretical and Applied Genetics 73, 809-814. Barwale, U.B. & Widholm, J.M. 1987 Somaclonal variation in plants regenerated from cultures of soybean. Plant Cell Reports 6, 365-368. Baxter, R., Brown, S.N., England, N.F., Ludlow, C.H.M., Taylor,

Biotechnology of trees S.L. & Womack, R.W. 1989 Production of clonal plantlets of tropical pine in tissue culture via axillary shoot activation. Canadian Journal of Forest Research 19, I338-1342. Beakbane, A.B. & Majumber, P.K. 1975 A relationship between stomatal density and growth potential in apple rootstocks. Journal of Horticultural Science 50, 285-289. Beakbane, A.B. & Thompson, E.C. 1939 Anatomical studies of stems and roots of hardy fruit trees. I. The internal structure of the roots of some vigorous and some dwarfing apple rootstocks and the correlation of structure with vigour. Journal of Pomology 17, 141-149. Bekkaoui, F., Pilon, M., Laine, E., Raju, D.S.S., Crosby, W.L. & Dunstan, D.I. I988 Transient gene expression in electroporated Picea glauca protoplasts. Plant Cell Reports 7, 481-484. Ben-Hayyim, G. & Goffer, Y. 1989 Plantlet regeneration from a NaCl-selected salt-tolerant callus culture of Shamouti orange (Citrus sinensis L. Osbeck). Plant Cell Reports 7, 680-683. Bon, M.-C. & Monteuuis, O. I991 Rejuvenation of a 100-year-old Sequoiadendron giganteum through in vitro culture. II. Biochemical arguments. Physiologia Plantarum 81, 116-120. Bravo, J.E. & Evans, D.A. 1985 Protoplast fusion for crop improvement. Plant Breeding Reviews 3, 193-218. Chambers, S.M., Heuch, J.H.R. & Pirrie, A. 1991 Micropropagation and in vitro flowering of the bamboo Dendrocalamus hamiltonii Munro. Plant Cell, Tissue and Organ Culture 27, 45-48. Comu, D. & Michel, M.F. 1987 Bacteria contaminants in shoot cultures of Prunus avium L. Choice and phytotoxicity of antibiotics. Acta Horticulturae 212, 83-86. De Block, M. I990 Factors influencing the tissue culture and the Agrobacterium tumefaciens-mediated transformation of hybrid aspen and poplar clones. Plant Physiology 93, 1110-1116. Donovan, A. 1991 Screening for fire blight resistance in apple (Malus pumila) using leaf assays from in vitro and in vivo grown material. Annals of Applied Biology 119, 59-68. Durzan, D.J. 1988 Rooting in woody perennials: problems and opportunities with somatic embryos and artificial seeds. Acta Horticulturae 227, 121-125. Fei, K.W. & Xue, G.R. 1981 Induction of haploid plantlets by anther culture in vitro in apple cv. Delicious. Scientia Agriculturae Sinica 4, 44. F6raud-Keller, C. & Espagnac, H. I989 Conditions d'apparition d'une embryog6n6se la culture de tissus foliaires du chine vert (Quercus ilex). Canadian Journal of Botany 67, I066-1070. Fillatti, J.J., Sellmer, J., McCown, B., Hassig, B. & Comai, L. 1987 Agrobacterium-mediated transformation and regeneration of Populus. Molecular and General Genetics 206, I92-199. Forestry Industry Committee of Great Britain 1987. Beyond 2000. London: FICGB. Gasser, C.S. & Fraley, R.T. 1989 Genetically engineering plants for crop improvement. Science 244, 1293-1299. Gr6nroos, L., von Arnold, S. & Eriksson, T. 1989 Callus production and somatic embryogenesis from floral explants of basket willow (Salix viminalis L.). Journal of Plant Physiology 134, 558-500. Grosser, J.W., Gmitter, F.G., Tusa, N. & Chandler, J.L. 1990 Somatic hybrid plants from sexually incompatible woody species: Citrus reticulata and Citropsis gilletiana. Plant Cell Reports 8, 656--059. Gupta, P.K., Dandekar, A.M. & Durzan, D.J. I988 Somatic proembryo formation and transient expression of a luciferase gene in Douglas Fir and Loblolly Pine protoplasts. Plant Science 58, 85-92. Hammatt, N. & Ridout, M.S. 1992 Micropropagation of common ash (Fraxinus excelsior). Plant Cell, Tissue and Organ Culture, in press. Hammerschlag, F.A. 1988 Selection of peach cells for insensitivity to culture filtrates of Xanthomonas campestris cv. pruni and

regeneration of resistant plants. Theoretical and Applied Genetics 76, 865-869. Hassig, B.E. 1989 Status of forest tree vegetative regeneration for biotechnology. American Biotechnology Laboratory 7, 48--5 I. Howard, B.H. 1987 Propagation. In Rootstocks for Fruit Crops, eds Rom, R.C. & Carlson, R.F. pp. 29-77. New York: Wiley. Howard, B.H., Jones, O.P. & Vasek, J. 1989 Long-term improvement in the rooting of plum cuttings following apparent rejuvenation. Journal of Horticultural Science 64, 147-156. James, D.J. 1987 Cell and tissue culture technology for the genetic manipulation of temperate fruit trees. In Biotechnology and Genetic Engineering Reviews, Vol. 5, ed Russell, G.E. pp. 33-79. Newcastle-upon-Tyne: Intercept. James, D.J., Mackenzie, K.A.D. & Malhotra, S.B. 1987 The induction of hexaploidy in cherry rootstocks using in vitro regeneration techniques. Theoretical and Applied Genetics 73, 589-594. James, D.J., Passey, A.J., Barbara, D.J. & Bevan, M. 1989 Genetic transformation of apple (,IVlalus pumila Mill.) using a disarmed Ti-binary vector. Plant Cell Reports 7, 658-061. Jones, O.P. 1992a Propagation in vitro of apple. In Micropropagation of Woody Plants, ed Ahuja, M.R. Dordrecht: Kluwer Academic Publishers. Jones, O.P. 1992b Field performance of micropropagated apple rootstocks. In Annual Report 1990--91. pp. 47-48. Wellesbourne, UK: Horticulture Research International. Jones, O.P. & Hadlow, W.C.C. 1989 Juvenile-like character of apple trees produced by grafting scions and rootstocks produced by micropropagation. Journal of Horticultural Science 64, 395-401. Jones, O.P. & Webster, C.A. I989 Improved rooting from conventional cuttings taken from micropropagated plants of Pyrus communis rootstocks. Journal of Horticultural Science 64, 429-434. Jones, O.P. & Vine, S.J. I908 The culture of gooseberry shoot tips for eliminating virus. Journal of Horticultural Science 43, 289-292. J6rgensen, J. 1988 Embryogenesis in Quercus petraea and Fagus sylvatica. Journal of Plant Physiology 132, 638-640. J6rgensen, J. 1989 Somatic embryogenesis in Aesculus hippocastahum L. by culture of filament callus. Journal of Plant Physiology 135, 240-241. Karp, A. I989 Can genetic instability be controlled in plant tissue cultures? IAPTC Newsletter 58, 2-11. KIimaszewska, K. I989 Recovery of somatic embryos and plantlets from protoplast cultures of Larix x eurolepis. Plant Cell Reports 8, 440--445. Kristiansen, K. I991 Post-propagation growth of cuttings from in vitro and in vivo propagated stock plants of Ficus benjamina. Scientia Horticulturae 46, 315-322. Lakshmi Sita, G., Raghava Ram, N.V. & Vaidyanathan, C.S. 1980 Triploid plants from endosperm cultures of sandalwood by experimental embryogenesis. Plant Science Letters 20, 63-69. Lapins, K.O. 1969 Segregation of compact growth types in certain apple seedling progenies. Canadian Journal of Plant Science 49, 765-768. Lapins, K.O. 1975 Polyploidy and mutations induced in apricot by colcbicine treatment. Canadian Journal of Genetics and Cytology 17, 591-599. Lapins, K.O. & Watkins, R. 1973 Genetics of compact growth habit. In Report of the East Mailing Research Station for 1972, p. 136. East Mailing, UK: East Malling Research Station. Lespinasse, Y., Godicheau, M. & Duron, M. 1983 Potential value and method of producing haploids in the apple tree Malus pumila (Mill.). Acta Horticulturae 131, 223-230. Lester, D.T. & Berbee, J.G. 1977 Within-clone variation among Black Poplar trees derived from callus culture. Forest Science 23, 122-131.

WorldJournalot Microbiologyand Biotechnology,VoI 8. 1992

3 75

N. Hammatt Levin, R. & Vasil, I.K. 1989 Progress in reducing the cost of micropropagation. IAPTC Newsletter 59, 1-12. Linington, I.M. 1991 In vitro propagation of Dipterocarpus alatus and Dipterocarpus intricatus. Plant Cell, Tissue and Organ Culture 27, 81-88. Mackenzie, K.A.D. 1989 Cambial activity and wood structure of micropropagated vs conventionally propagated trees. In Annual Report of the AFRC Institute of Horticultural Research for 1988, pp. 32-33. East Mailing, UK: Agricultural and Food Research Council. Manganaris, A.G. & Alston, F.H. 1987 Inheritance and linkage relationships of glutamate oxaloacetate transarninase isoenzymes in apple. I. The gene GOT-I, a marker for the S incomparability locus. Theoretical and Applied Genetics 74, 154-161. Mante, S., Morgens, P.H., Scorza, R., Cordts, J.M. & Callahan, A.M. 1991 Agrobacterium-mediated transformation of plum (Prunus domestica L.) hypocotyl slices and regeneration of transgenic plants. Bio/Technology 9, 853-857. Manzanera, J.A. & Pardos, J.A. 1990 Micropropagation of juvenile and adult Quercus suber L. Plant Cell, Tissue and Organ Culture 21, 1-8. Marks, T.R. & Simpson, S.E. 1990 Reduced phenolic oxidation at culture initiation in vitro following the exposure of field-grown stockplants to darkness or low levels of irradiance. Journal of Horticultural Science 65, 103-111. McGranahan, G.H., Leslie, C.A., Uratsu, S.L., Martin, L.A. & Dandekar, A.M. 1988 Agrobacterium-mediated transformation of walnut somatic embryos and regeneration of transgenic plants. Bio/Technology 6, 800-804. Nagauda, R.S., Parasharami, V.A. & Mascarenhas, A.F. 1990 Precocious flowering and seedling behaviour in tissue-cultured bamboos. Nature 344, 335-336. Navatel, J.C., Nio, M., Vaysse, P. & Edin, M. 1988 Pommier. Comportement en marcotti6res et aux vergers des porte-greffes micropropag6s de type Malus. Infos-Ctifl 4, 25-28. Ochatt, S.J., Cocking, E.C. & Power, J.B. 1987 Isolation, culture and plant regeneration of colt cherry (Prunus avium x pseudocerasus) protoplasts. Plant Science 50, 139-143. Ochatt, S.J., Patat-Ochatt, E.M., Reck E.L., Davey, M.R. & Power, J.B. 1989 Somatic hybridization of sexually incompatible top-fruit tree rootstocks, wild pear (Pyrus communis var. pyraster L.) and Colt cherry (Prunus avium x pseudocerasus). Theoretical and Applied Genetics 78, 35-41. Ochatt, S.J. & Power, J.B. 1988a Plant regeneration from mesophyll protoplasts of William's Bon Chretien (syn Bartlett) pear (Pyrus communis L.). Plant Cell Reports 7, 587-589. Ochatt, S.J. & Power, J.B. 1988b An alternative approach to plant regeneration from protoplasts of sour cherry (Prunus cerasus L.). Plant Science 56, 75-79. Ochatt, S.J. & Power, J.B. 1989 Selection for salt and drought tolerance in protoplast- and explant-derived tissue cultures of Colt cherry (Prunus avium x pseudocerasus). Tree Physiology 5, 259-266. Ostry, M.E. & Skilling, D.D. 1988 Somatic variation in resistance of Populus to Septoria musiva. Plant Disease 72, 724-727. Patat-Ochatt, E.M., Ochatt, S.J. & Power, J.B. 1988 Plant regeneration from protoplasts of apple rootstocks and scion varieties (Malus x domestica Borklh.). Journal of Plant Physiology 133, 460--465. P6rinet, P., Vall6e, G. & Tremblay, F.M. 1988 In vitro propagation of mature trees of Alnus incana (L.) Moench. Plant Cell, Tissue and Organ Culture 15, 85-89. Phelep, M., Petit, A., Martin, L., Duhoux, E. & Temp6, J. 1991 Transformation and regeneration of a nitrogen-fixing tree,

3 76

WorldJournalof Microbiolog~/andBiotechnology,Vol 8, 1992

Allocasuarina verticillata Lam. Bio/Technology 9, 461-466. Prakash, C.S. & Thielges, B.A. 1989 Somaclonal variation in Eastern Cottonwood for race-specified partial resistance to leaf rust disease. Phytopathology 79, 805-808. Preece, J.E., Huetteman, C.A., Ashby, W.C. & Roth, P.L. I991a Micro- and cutting propagation of silver maple. I Results with adult and juvenile propagules. Journal of the American Society for Horticultural Science 116, 142-148. Preece, J.E., Huetteman, C.A., Ashby, W.C. & Roth, P.L. 1991b Micro- and cutting propagation of silver maple. Il Genotype and provenance affect performance. Journal of the American Society for Horticultural Science 116, 149-155. Preece, J.E., Zhao, J. & Kung, F.H. 1989 Callus production and somatic embryogenesis from White Ash. HortScience 24, 377-380. Quinlan, J.D. & Tobutt, K.R. 1990 Manipulating fruit tree structure chemically and genetically for improved performance. HortScience 25, 60--64. Radojevi6, L. 1978 In vitro induction of androgenic plantlets in Aesculus hippocastanum L. Protoplasma 96, 369-374. Radojevi6, L., Djordjevih, N. & Tuci4, B. 1989 In vitro induction of pollen embryos and plantlets in Aesculus carnea Hayne through anther culture. Plant Cell, Tissue and Organ Culture 17, 21-26. Radojevih, L. & Kovoor, A. 1985 Induction of haploids. In Biotechnology in Agriculture and Forestry. I, Trees I, ed Bajaj, Y.P.S. pp. 65-68 Berlin: Springer Verlag. Rugini, E. & Fontanazza, G. 1981 In vitro propagation of "Dolce Agogia" olive. HortScience 16, 492-493. Russell, J.A. & McCown, B.H. 1988 Recovery of plants from leaf protoplasts of hybrid-poplar and aspen clones. Plant Cell Reports 7, 59--62. S6guin, A. & Lalonde, M. 1988 Gene transfer by electroporation in Betulaceae protoplasts: Alnus incana. Plant Cell Reports 7, 367-370. Sim, F.E., Loh, C.J. & Goh, C.J. 1988 Direct somatic embryogenesis from protoplasts of Citrus mitis Blanco. Plant Cell Reports 7, 418-420. Smigocki, A.C. & Hammerschlag, F.A. 1991 Regeneration of plants from peach embryo cells infected with a shooty mutant strain of Agrobacterium. Journal of the American Society for Horticultural Science 116, 1092-1097. Sticklen, M.B., Domir, S.C. & Lineberger, R.D. 1986 Shoot regeneration from protoplasts of Ulmus x 'Pioneer'. Plant Science 47, 29-34. Teasdale, R.D. & Buxton, P.A. 1986 Culture of Pinus radiata embryos with reference to artificial seed production. New Zealand Journal of Forest Science 16, 387-391. Tobutt, K.R. 1985 Breeding columnar apples at East Malling. Acta Horticulturae 189, 63--68. Tulecke, W., McGranahan, G. & Ahmadi, H. 1988 Regeneration by somatic embryogenesis of triploid plants from endosperm of walnut, ]uglans regia L. cv Manregian. Plant Cell Reports 7, 301-304. Tydeman, H.M. & Alston, F.H. 1965 The influence of dwarfing rootstocks in shortening the juvenile phase of apple seedlings. In Report of the East Mailing Research Station for I964, pp. 97-98. East Mailing, UK: East Mailing Research Station. Valobra, C.P. & James, D.J. 1990 In vitro shoot regeneration from leaf discs of Betula pendula 'Dalecarlica' EM 85. Plant Cell, Tissue and Organ Culture 21, 51-54. Vardi, A., Bleichman, S. & Aviv, D. 1990 Genetic transformation of Citrus protoplasts and regeneration of transgenic plants. Plant Science 69, 199-206. Vardi, A, Breiman, A. & Galun, E. 1987 Citrus cybrids: production

Biotechnology of trees by donor-recipient protoplast-fusion and verification by mitochondrial-DNA restriction profiles. Theoretical and Applied Genetics 75, 51-58. Vardi, A., Spiegel-Roy, P. & Galun, E. 1982 Plant regeneration from Citrus protoplasts: variability in methodological requirements among cultivars and species. Theoretical and Applied Genetics 62, 171-176. Vieitez, A.M. & Vieitez, M.L. 1980 Culture of chestnut shoots from buds in vitro. Journal of Horticultural Science 55, 83-84. Von Arnold, S. & Hakman, I. 1988 Regulation of somatic embryo development in Picea abies by abscisic acid. Journal of Plant Physiology 132, 164-169+ Wang, T.-Y. & Chang, C.-J. 1978 Triploid Citrus plantlet from endosperm culture. Scientia Sinica 21, 823-827. Webster, C.A. & Jones, O.P. 1989a Micropropagation of the apple rootstock M.9: effect of sustained subculture on apparent rejuvenation in vitro. Journal of Horticultural Science 64, 421-428. Webster, C.A. & Jones, O.P. I989b Effects of sustained subculture on apparent rejuvenation of the apple rootstock M.9 in vitro and in vivo. Annales des Sciences Foresti&es 46, 187S-189S.

Webster, A.D., Sparks, T.R. & Blecher, A. 1986 The influence of micropropagation and chemical mutagens on the growth and precocity of Cox's Orange Pippin and Bramley's Seedling apple. Acta Horticulturae 180, 25-34. Wilson, Z.A. & Power, J.B. 1989 Elimination of systemic contamination in explant and protoplast cultures of rubber (Hevea brasiliensis Muell. Arg.). Plant Cell Reports 7, 622-625. Wilson, S.M., Thorpe, T.A. & Moloney, M.M. 1989 PEG-mediated expression of GUS and CAT genes in protoplasts from embryogenic suspension cultures of Picea glauca. Plant Cell Reports 7, 707-709. Wu, J. 1982 Obtaining haploid plantlets of crab apple from anther culture in vitro. Acta Horticulturae Sinica 8, 36. Zimmerman, R.H. & Miller, S.S. 1991 Orchard growth and fruiting of micropropagated apple trees. Journal of the American Society for Horticultural Science 116, 780-785. (Received in revised form 14 February I992, accepted I4 February 1992)

Worm Journal of Mzcrobiotogy and BiotechnoIogy, Vo[ 8, 1992

377

Progress in the biotechnology of trees.

An increasing world population and rise in demand for tree products, especially wood, has increased the need to produce more timber through planting m...
904KB Sizes 2 Downloads 0 Views