Agrobacterium tumefaciens and related Agrobacterium species have been known as plant pathogens since the beginning of the 20th century. However, only in the past two decades has the ability of Agrobacterium to transfer DNA to plant cells been harnessed for the purposes of plant genetic engineering. Since the initial reports in the early 1980s using Agrobacterium to generate transgenic plants, scientists have attempted to improve this “natural genetic engineer” for biotechnology purposes. Some of these modifications have resulted in extending the host range of the bacterium to economically important crop species. However, in most instances, major improvements involved alterations in plant tissue culture transformation and regeneration conditions rather than manipulation of bacterial or host genes. Agrobacterium-mediated plant transformation is a highly complex and evolved process involving genetic determinants of both the bacterium and the host plant cell. In this article, I review some of the basic biology concerned with Agrobacterium-mediated genetic transformation. Knowledge of fundamental biological principles embracing both the host and the pathogen have been and will continue to be key to extending the utility of Agrobacterium for genetic engineering purposes.

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Agrobacterium-Mediated Plant Transformation: the Biology behind the “Gene-Jockeying” Tool

Section A 1 Somatic Embryogenesis in White Spruce: Studies of Embryo Development and Cell Biology L Kong, et al 2 Proliferative Somatic Embryogenesis in Woody Species K Raemakers, et al 3 Somatic Embryo Germination and Desiccation Tolerance in Conifers EI Hay, PJ Charest 4 Performance of Conifer Stock Produced Through Somatic Embryogenesis SC Grossnickle 5 Apoptosis During Early Somatic Embryogenesis in Picea spp L Havel, DJ Durzan 6 Water Relation Parameters in Conifer Embryos: Methods and Results N Dumont-Beboux, et al 7 Image Analysis for Sorting Somatic Embryos Y Ibaraki 8 Somatic Embryogenesis in Woody Legumes RN Trigiano, et al 9 Cold Storage and Cryopreservation of Camellia Embryogenic Cultures AM Vieitez, A Ballester 10 Cryopreservation of Embryogenic Cultures of Conifers and Its Application to Clonal Forestry DR Cyr 11 Commercialization of Plant Somatic Embryogenesis BCS Sutton, DR Polonenko Section B 12 Somatic Embryogenesis in Myrtaceous Plants JM Canhoto, et al 13 Somatic Embryogenesis Induction in Bay Laurel (Laurus nobilis L) JM Canhoto, et al 14 Somatic Embryogenesis in Simarouba glauca Linn GR Rout, P Das 15 Somatic Embryogenesis in Magnolia spp SA Merkle 16 Somatic Embryogenesis and Evaluation of Variability in Somatic Seedlings of Quercus serata by RAPD Markers K Ishii, et al 17 Somatic Embryogenesis from Immature Fruit of Juglans cinerea PM Pijut Section C 18 Somatic Embrygonesis in Pinus patula Scheide et Deppe NB Jones, J van Staden 19 Somatic Embryogenesis in African Cycads (Encephalartos) AK Jager, J van Staden 20 Somatic Embryogenesis in Picea wilsonii Y Yang, Z Guo 21 Somatic Embryogenesis in Jack Pine (Pinus banksiana Lamb) YS Park, et al 22 Somatic Embryogenesis in Hybrid Firs J Jasik, et al 23 Somatic Embryogenesis in Taxus SR Wann, et al

Somatic embryogenesis in woody plants

Terpenoid biomaterials

The role of activated charcoal in plant tissue culture.

transgenic plant overexpressing a recombinant polynucleotide, wherein said transgenic plant has improved compared production with a non-transgenic plant or wild type plant, wherein the polynucleotide encodes recombinant encodes a polypeptide having at least 95% identity in amino acid sequence over the entire length of SEQ ID nO: 328, said polypeptide providing improved compared to a non-transgenic plant that does not overexpress the polypeptide production.

Polynucleotides and polypeptides in plants

The effects of different concentrations of hydroxyurea (HU) and aphidicolin (APH) on the mitotic index (MI) were compared in cells of embryogenic cultures of Larix decidua, L. leptolepis, and L. decidua x L. leptolepis (Larix x eurolepis). The highest enhancement of the MI was obtained with HU at 1.25 mM and 0.6% colchicine. In general the MI decreased with an increase of HU or APH concentration (over 1.25 mM for HU and 5 μM for APH). Detailed karyotype analyses were made on the somatic complement of L. decidua, L. leptolepis, and their hybrid. These karyotypes were asymmetrical and advanced, with the smaller chromosomes being more submedian than the larger ones. The topography of chromosome 7 of L. decidua and chromosome 9 of L. leptolepis was found to be the most significant cytotaxonomic characteristic in differentiating these two species. Cytological data indicate that Japanese larch (L. leptolepis) is phylogenetically closer to European larch (L. decidua) than the Siberian larch group (L. sibirica and L. sukaczewii). Chromosomes with unusually long kinetochores were found in both species and the hybrid. Hyperploid cells (2n = 25) were observed in the hybrid (Larix x eurolepis) material analyzed. A genomic L. decidua probe hybridized strongly to dots of DNA from L. leptolepis indicating that there is high sequence homology between these two species.

Cytological and molecular relationships between Larix decidua, L. leptolepis and Larix x eurolepis: identification of species-specific Chromosoms and synchronization of mitotic cells.

Somatic embryogenesis (SE) of conifers for clonal propagation has emerged from its earliest beginnings in 1980 to become an integral component of tree improvement strategies. With its capacity for long-term germplasm preservation and scale-up technologies, it is seen to be the preferred avenue to accelerate selection and operational deployment of value-added genotypes. At present, programs for numerous species from the Pinaceae family are underway worldwide, with activities ranging from selection trials to pilot production for plantation forestry. This paper will provide a review of current efforts in conifer SE including cryopreservation, commercialization and deployment strategies, and transgenics. A discussion of challenges and issues is directed at genetic fidelity, intellectual property and future needs. Additional key words: clonal propagation, germplasm preservation, Pinaceae, plantation forestry, somatic

http://agro.icm.edu.pl/agro/element/bwmeta1.element.agro-article-e0b76308-4acd-48c3-9d2d-193a4de72f27/c/48_41_49.pdf

Conifer somatic embryogenesis: II. Applications

Global transition towards a bioeconomy sets new demands for wood supply (bioenergy, biomaterials, biochemicals, etc.), and the forestry sector is also expected to help mitigate climate change by increasing carbon fixation. For increased biomass production, the use of improved, genetically superior materials becomes a necessity, and vegetative propagation of elite genotypes provides a potential delivery mechanism for this. Vegetative propagation through somatic embryogenesis alone or in combination with rooted cuttings obtained from somatic young trees can facilitate both tree breeding (greater selection accuracy and gains, breeding archives of donor material for making crosses after selection) and the implementation of deployment strategies for improved reforestation materials. To achieve these goals, progress in the efficiency of pine somatic embryogenesis biotechnology has been made for a few commercial pine species, and a better understanding has been gained of the molecular mechanisms underpinning somatic and zygotic embryo development.

Somatic Embryogenesis for More Effective Breeding and Deployment of Improved Varieties in Pinus spp.: Bottlenecks and Recent Advances

Among the many commercial opportunities afforded by somatic embryogenesis (SE), it is the ability to clonally propagate individual plants with rare or elite traits that has some of the most significant implications. This is particularly true for many long-lived species, such as conifers, but whose long generation times pose substantive challenges, including increased recalcitrance for SE as plants age. Identification of a clonal line of somatic embryo-derived trees whose shoot primordia have remained responsive to SE induction for over a decade, provided a unique opportunity to examine the molecular aspects underpinning SE within shoot tissues of adult white spruce trees. Microarray analysis was used to conduct transcriptome-wide expression profiling of shoot explants taken from this responsive genotype following one week of SE induction, which when compared with that of a nonresponsive genotype, led to the identification of four of the most differentially expressed genes within each genotype. Using absolute qPCR to expand the analysis to three weeks of induction revealed that differential expression of all eight candidate genes was maintained to the end of the induction treatment, albeit to differing degrees. Most striking was that both the magnitude and duration of candidate gene expression within the nonresponsive genotype was indicative of an intense physiological response. Examining their putative identities further revealed that all four encoded for proteins with similarity to angiosperm proteins known to play prominent roles in biotic defense, and that their high-level induction over an extended period is consistent with activation of a biotic defense response. In contrast, the more temperate response within the responsive genotype, including induction of a conifer-specific dehydrin, is more consistent with elicitation of an adaptive stress response. While additional evidence is required to definitively establish an association between SE responsiveness and a specific physiological response, these results suggest that biotic defense activation may be antagonistic, likely related to the massive transcriptional and metabolic reprogramming that it elicits. A major issue for future work will be to determine how and if suppressing biotic defense activation could be used to promote a physiological state more conducive to SE induction.

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Potential link between biotic defense activation and recalcitrance to induction of somatic embryogenesis in shoot primordia from adult trees of white spruce (Picea glauca)

Over the last 5 yr, the production of transgenic conifers has been greatly facilitated by the ability to transform somatic embryonal tissues (somatic embryos) via cocultivation with Agrobacterium tumefaciens. This has allowed us to develop protocols for the genetic transformation of several spruce species. Furthermore, these procedures can produce an average of 20 independent transgenic lines (translines) per gram fresh mass of embryonal tissue, providing for the first time the magnitude-of-scale required for implementing large-scale functional genomics studies in conifers. Combined with efficient regeneration of transgenic trees via somatic embryos, the potential for genetic engineering of conifers has been demonstrated by stable reporter gene expression (GUS or GFP) resulting from single insert T-DNA integration events.

Krystyna Klimaszewska

Papers

Cytological and molecular relationships between Larix decidua, L. leptolepis and Larix x eurolepis: identification of species-specific Chromosoms and synchronization of mitotic cells.

Conifer somatic embryogenesis: II. Applications

Somatic Embryogenesis for More Effective Breeding and Deployment of Improved Varieties in Pinus spp.: Bottlenecks and Recent Advances

Potential link between biotic defense activation and recalcitrance to induction of somatic embryogenesis in shoot primordia from adult trees of white spruce (Picea glauca)

Genetic Transformation of Conifers Utilizing Somatic Embryogenesis