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The genus Quercus, which belongs to the family Fagaceae, is native to the northern hemisphere and includes deciduous and evergreen species. The trees of the different species are very important from both economic and ecological perspectives. Application of new technological approaches (which span the fields of plant developmental biology, genetic transformation, conservation of elite germplasm and discovery of genes associated with complex multigenic traits) to these long-rotation hardwoods may be of interest for accelerating tree improvement programs. This review provides a summary of the advances made in the application of biotechnological tools to specific oak species. Significant progress has been made in the area of clonal propagation via organogenesis and somatic embryogenesis (SE). Standardized procedures have been developed for micropropagating the most important European (Q. robur, Q. petarea, Q. suber) and American (Q. alba, Q. bicolor, Q. rubra) oaks by axillary shoot growth. Although regenerated plantlets are grown in experimental trials, large-scale propagation of oak species has not been carried out. The induction of SE in oaks from juvenile explants is generally not problematic, although the use of explants other than zygotic embryos is much less efficient. During the last decade, enormous advances have been made in inducing SE from selected adult trees, mainly specimens of pedunculate oak (Q. robur) and cork oak (Q. suber). Advances in the understanding of the maturation and germination steps are required for better use of embryogenic process in clonal forestry. Quercus species are late-maturing and late-flowering, exhibit irregular seed set, and produce seeds that are recalcitrant to storage by conventional procedures. Vitrification-based cryopreservation techniques were used successfully in somatic embryos of pedunculate oak and cork oak, and an applied genbank of cork oak selected genotypes is now under development. The feasibility of genetic transformation of pedunculate oak and cork oak somatic embryos by means of co-culture techniques with several strains of Agrobacterium tumefaciens has also been demonstrated. To date, most research on the genomics of Quercus species has concerned population genetics. Approaches using functional genomics to examine the molecular and cellular mechanisms that control organogenesis and or somatic embryogenesis are still scarce, and efforts on the isolation and characterization of genes related to other specific traits should be intensified in the near future, as this would help improve the practical application of clonal forestry in recalcitrant species such as oaks.

In-vitro-cultured shoots of clones initiated from shoots of the basal parts (BS) and the crown (C) of two mature Quercus robur L. trees were subjected to rooting experiments to relate rooting with shoot topophysical origin. The BS shoots exhibited morphologically juvenile characteristics and rooted more easily after indole-3-butyric acid (IBA) treatment than C shoots. When naphthylphthalamic acid (NPA) was applied to BS shoots, rooting capacity decreased and root emergence was delayed at least 2 days compared with shoots treated with IBA only. During the first days of the rooting process, endogenous indole-3-acetic acid (IAA) concentration was higher in C shoots than in BS shoots, regardless of whether the shoots were treated with NPA. Mitotic figures were observed in cells from the basal part of both BS and C shoots 24 h after IBA treatment. After 4 days of IBA treatment, the occurrence of histological events differed between BS shoots and C shoots. Cells of BS shoots became meristematic, giving rise to meristemoids and root primordia, whereas no differentiation of root meristemoids occurred in cells of C shoots. Thus, although adult oak material (C shoots) is capable of responding to the initial stimulus of auxin during the adventitious rooting process, the endogenous IAA concentration is not the factor limiting rooting in adult material.

A protocol for culturing chestnut axillary shoots by temporary immersion in liquid medium was developed. The influence of type of explant, support material, bioreactor, and immersion was investigated for five artificial hybrids and five natural hybrids of Asian and European chestnut selected for resistance to ink disease. The type of explant influenced shoot quality and proliferation rates, and basal explants with callus produced more and longer shoots than apical and nodal segments. Use of rockwool cubes as support material prevented hyperhydricity and allowed proliferation of explants in Murashige and Skoog medium with half-strength nitrates supplemented with 0.22 lM BA and 3 % sucrose, cultured both in plantform TM and RITA Ò vessels with three or six immersions per day and additional aeration of 1 min per hour in the case of plantform TM bioreactors. Basal explants cultured in plantform TM for 5 weeks produced long shoots suitable for rooting, whereas apical and nodal explants cultured in plantform TM or RITA Ò produced shorter shoots that were suitable for maintenance of stock. For most of the clones, similar or higher proliferation rates were observed when cultured in liquid medium than in semisolid medium, with the additional benefit of cost-reduction of the former system. Shoots developed in liquid medium were submitted to ex vitro root induction by dipping in indole-3-butyric acid, and acclimatized under greenhouse conditions. This is the first demonstration of the production of chestnut plantlets from shoots cultured in liquid medium, and the protocol presented here shows good potential for application in large-scale propagation.

Plant biotechnology can be used to conserve the germplasm of natural forests, and to increase the productivity and sustainability of plantations. Both goals imply working with mature trees, which are often recalcitrant to micropropagation. Conventional in vitro culture uses closed containers and gelled medium with sugar supplementation. Bioreactor culture uses liquid medium and usually incorporates aeration. The increased absorption of nutrients via the liquid medium together with the renewal of the air inside the bioreactors may improve the physiological state of the explants. In this review, we will explore the feasibility of using bioreactors to overcome the recalcitrance of many trees to micropropagation and/or to decrease the cost of large‐scale propagation. We will focus on the recent use of bioreactors during the multiplication, rooting (plant conversion in the case of somatic embryos), and acclimation stages of the micropropagation of axillary shoots and somatic embryos of forest trees (including some shrubs of commercial interest), in both temporary and continuous immersion systems. We will discuss the advantages and the main obstacles limiting the widespread implementation of bioreactor systems in woody plant culture, considering published scientific reports and contributions from the business sector.

The genus Castanea includes several tree species that are relevant because of their geographical extension and their multipurpose character, that includes nut and timber production. However, commercial exploitation of the trees is hindered by several factors, particularly by their limited regeneration ability. Regardless of recent advances, there exists a serious limitation for the propagation of elite genotypes of chestnut due to decline of rooting ability as the tree ages. In the present review, we summarize the research developed in this genus during the last three decades concerning the formation of adventitious roots (ARs). Focusing on cuttings and in vitro microshoots, we gather the information available on several species, particularly C. sativa, C. dentata and the hybrid C.sativa × C. crenata, and analyze the influence of several factors on the achievements of the applied protocols, including genotype, auxin treatment, light regime and rooting media. We also pay attention to the acclimation phase, as well as compile the information available about biochemical and molecular related aspects. Furthermore, we considerate promising biotechnological approaches that might enable the improvement of the current protocols.

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