Horizontal gene transfer contributes to plant evolution : the case of Agrobacterium T-DNAs

Horizontal gene transfer (HGT) can be defined as the acquisition of genetic material from another organism without being its offspring. HGT is common in the microbial world including archaea and bacteria, where HGT mechanisms are widely understood and recognized as an important force in evolution. In eukaryotes, HGT now appears to occur more frequently than originally thought. Many studies are currently detecting novel HGT events among distinct lineages using next-generation sequencing. Most examples to date include gene transfers from bacterial donors to recipient organisms including fungi, plants, and animals. In plants, one well-studied example of HGT is the transfer of the tumor-inducing genes (T-DNAs) from some Agrobacterium species into their host plant genomes. Evidence of T-DNAs from Agrobacterium spp. into plant genomes, and their subsequent maintenance in the germline, has been reported in Nicotiana, Linaria and, more recently, in Ipomoea species. The transferred genes do not produce the usual disease phenotype, and appear to have a role in evolution of these plants. In this paper, we review previous reported cases of HGT from Agrobacterium, including the transfer of T-DNA regions from Agrobacterium spp. to the sweetpotato [ Ipomoea batatas (L.) Lam.] genome which is, to date, the sole documented example of a naturally-occurring incidence of HGT from Agrobacterium to a domesticated crop plant. We also discuss the possible evolutionary impact of T-DNA acquisition on plants

Optical microscopy on agrobacterium-mediated transient transformated arabidopsis nahg plants

Agrobacterium tumefaciens-mediated transient transformation has demonstrated to be an invaluable tool in plant cell biology. However, low efficiency and inconsistency of this method in Arabidopsis has forced the implementation of Nicotiana benthamiana as a surrogate system, limiting applicability. Previous results suggested that hormone-mediated defence responses against bacteria might be responsible for the low efficiency of Agrobacterium-mediated transient transformation in Arabidopsis. In this work, we evaluate the efficiency of Agrobacterium-mediated transient transformation in Arabidopsis genotypes affected in JA perception or signalling (coi1, jin1), or with low SA or JA content (sid2, NahG, aos). We demonstrate that expression of the NahG transgene dramatically improves this process. Arabidopsis NahG plants can be efficiently used for transient expression-based optical microscopy assays routinely performed in N. benthamiana, such as determination of subcellular localization of GFP-fused proteins or analysis of protein-protein interactions by Bimolecular Fluorescent Complementation. Considering the wide-spread use of Agrobacterium-mediated transient transformation, this system can enormously facilitate research in the model plant Arabidopsis, allowing for an efficient use of the full potential of the numerous tools and resources currently available to the community.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Enhancement of Recombinant Protein Production in Transgenic Nicotiana benthamiana Plant Cell Suspension Cultures with Co-Cultivation of Agrobacterium Containing Silencing Suppressors.

We have previously demonstrated that the inducible plant viral vector (CMViva) in transgenic plant cell cultures can significantly improve the productivity of extracellular functional recombinant human alpha-1-antiryspin (rAAT) compared with either a common plant constitutive promoter (Cauliflower mosaic virus (CaMV) 35S) or a chemically inducible promoter (estrogen receptor-based XVE) system. For a transgenic plant host system, however, viral or transgene-induced post-transcriptional gene silencing (PTGS) has been identified as a host response mechanism that may dramatically reduce the expression of a foreign gene. Previous studies have suggested that viral gene silencing suppressors encoded by a virus can block or interfere with the pathways of transgene-induced PTGS in plant cells. In this study, the capability of nine different viral gene silencing suppressors were evaluated for improving the production of rAAT protein in transgenic plant cell cultures (CMViva, XVE or 35S system) using an Agrobacterium-mediated transient expression co-cultivation process in which transgenic plant cells and recombinant Agrobacterium carrying the viral gene silencing suppressor were grown together in suspension cultures. Through the co-cultivation process, the impacts of gene silencing suppressors on the rAAT production were elucidated, and promising gene silencing suppressors were identified. Furthermore, the combinations of gene silencing suppressors were optimized using design of experiments methodology. The results have shown that in transgenic CMViva cell cultures, the functional rAAT as a percentage of total soluble protein is increased 5.7 fold with the expression of P19, and 17.2 fold with the co-expression of CP, P19 and P24

Acquisition of an Agrobacterium Ri Plasmid and Pathogenicity by Other -Proteobacteria in Cucumber and Tomato Crops Affected by Root Mat

Root mat of cucumbers and tomatoes has previously been shown to be caused by Agrobacterium radiobacter strains harboring a root-inducing Ri plasmid (pRi). Nine other pRi-harboring -Proteobacteria have subsequently been isolated from root mat-infected crops. Fatty acid profiling and partial 16S rRNA sequence analysis identified three of these strains as being in the genus Ochrobactrum, five as being in the genus Rhizobium, and one as being in the genus Sinorhizobium. An in vitro pathogenicity test involving inoculation of cucumber cotyledons was developed. All pRi-harboring -Proteobacteria induced typical root mat symptoms from the cotyledons. Average transformation rates for rhizogenic Ochrobactrum (46%) and Rhizobium (44%) strains were lower than those observed for rhizogenic A. radiobacter strains (64%). However, individual strains from these three genera all had transformation rates comparable to those observed from cotyledons inoculatedwith a rhizogenic Sinorhizobium strain (75%)

Phylogenies of atpD and recA support the small subunit rRNA-based classification of rhizobia

The current classification of the rhizobia (root-nodule symbionts) assigns them to six genera. It is strongly influenced by the small subunit (16S, SSU) rRNA molecular phylogeny, but such single-gene phylogenies may not reflect the evolution of the genome as a whole. To test this, parts of the atpD and recA genes have been sequenced for 25 type strains within the alpha -Proteobacteria, representing species in Rhizobium, Sinorhizobium, Mesorhizobium, Bradyrhizobium, Azorhizobium, Agrobacterium, Phyllobacterium, Mycoplana and Brevundimonas. The current genera Sinorhizobium and Mesorhizobium are well supported by these genes, each forming a distinct phylogenetic clade with unequivocal bootstrap support. There is good support for a Rhizobium clade that includes Agrobacterium tumefaciens, and the very close relationship between Agrobacterium rhizogenes and Rhizobium tropici is confirmed. There is evidence for recombination within the genera Mesorhizobium and Sinorhizobium, but the congruence of the phylogenies at higher levels indicates that the genera are genetically isolated. rRNA provides a reliable distinction between genera, but genetic relationships within a genus may be disturbed by recombination

Plant Cellular and Molecular Biotechnology: Following Mariotti's Steps

This review is dedicated to the memory of Prof. Domenico Mariotti, who significantly contributed to establishing the Italian research community in Agricultural Genetics and carried out the first experiments of Agrobacterium-mediated plant genetic transformation and regeneration in Italy during the 1980s. Following his scientific interests as guiding principles, this review summarizes the recent advances obtained in plant biotechnology and fundamental research aiming to: (i) Exploit in vitro plant cell and tissue cultures to induce genetic variability and to produce useful metabolites; (ii) gain new insights into the biochemical function of Agrobacterium rhizogenes rol genes and their application to metabolite production, fruit tree transformation, and reverse genetics; (iii) improve genetic transformation in legume species, most of them recalcitrant to regeneration; (iv) untangle the potential of KNOTTED1-like homeobox (KNOX) transcription factors in plant morphogenesis as key regulators of hormonal homeostasis; and (v) elucidate the molecular mechanisms of the transition from juvenility to the adult phase in Prunus tree species

U.S. Patent Literature Survey of Agrobacterium-Mediated Transformation of Sweet Potato (Ipomoea Batatas)

A team of researchers and patent information scientists at Franklin Pierce Law Center were asked to evaluate the patent and literature landscape related to the Agrobacterium-mediated transformation in sweet potato with respect to the U.S. patents and patent applications. This report provides a patent landscape of the Agrobacterium-mediated transformation of sweet potato. The report includes the applicable methods of transformation and has also included certain patents and patent applications which claim a transformed plant by virtue of these methods. In certain cases, the claim structure covers Agrobacterium-mediated transformation technology via system and composition of matter claims and not the more prevalent method claims. Sweet potato plant (Ipomoea batatas) is adaptable to a broad range of agroecological conditions and fits in low input agriculture. It is highly productive even under adverse farming conditions. Sweet potato is grown in more than 100 countries as a valuable source of food, animal feed and industrial raw material. It is a staple crop in many South East Asian and African countries. Traditional plant breeding has contributed to the improvement of sweet potato, especially in developed countries such as the U.S.A. and Japan. Because of the biological complexities of sweet potato, sexual hybridization strategies have not been very effective in developing improved cultivars. Confidential Therefore, biotechnological tools, such as gene transfer, are very attractive in sweet potato improvement, as they enable direct introduction of desirable genes from other sources into preadapted cultivars

Cyclic β-glucans at the bacteria–host cells interphase: One sugar ring to rule them all

Cyclic β‐1,2‐D‐glucans (CβG) are natural bionanopolymers present in the periplasmic space of many Proteobacteria. These molecules are sugar rings made of 17 to 25 D‐glucose units linked exclusively by β‐1,2‐glycosidic bonds. CβG are important for environmental sensing and osmoadaptation in bacteria, but most importantly, they play key roles in complex host–cell interactions such as symbiosis, pathogenesis, and immunomodulation. In the last years, the identification and characterisation of the enzymes involved in the synthesis of CβG allowed to know in detail the steps necessary for the formation of these sugar rings. Due to its peculiar structure, CβG can complex large hydrophobic molecules, a feature possibly related to its function in the interaction with the host. The capabilities of the CβG to function as molecular boxes and to solubilise hydrophobic compounds are attractive for application in the development of drugs, in food industry, nanotechnology, and chemistry. More importantly, its excellent immunomodulatory properties led to the proposal of CβG as a new class of adjuvants for vaccine development.Fil: Guidolin, Leticia Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Arce Gorvel, Vilma. Centre National de la Recherche Scientifique; FranciaFil: Ciocchini, Andres Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Comerci, Diego José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; Argentina. Comisión Nacional de Energía Atómica. Centro Atómico Ezeiza; ArgentinaFil: Gorvel, Jean-Pierre. Centre National de la Recherche Scientifique; Franci

Regeneration ability and genetic transformation of root type chicory (Cichorium intybus var. sativum)

To develop an efficient protocol for shoot regeneration of root chicory (Cichorium intybus var. sativum), some factors, including different concentrations of plant growth regulators in Murashige and Skoog (MS) medium, type of explants and genotypes were evaluated. Initiation of callusing were best achieved in MS medium supplemented with 1-naphthaleneacetic acid (NAA) (0.1 mg l-1) plus 6-Benzylaminopurine (6-BAP) (1 mg l-1), indole-3-acetic acid (IAA) (0.01 mg l-1) plus 6-BAP (1.0 mg l-1), and IAA (0.5 mg l-1) plus (0.5 mg l-1) 6-BAP combinations on leaf and cotyledon explants. Explant-derived calli were able to produce multiple adventitious shoots in MS medium containing IAA (0.5 mg l-1) plus 6-BAP (0.5 mg l-1). MS medium containing indole-3-butylric acid IBA (1 mgl-1) efficiently induced rooting on elongated shoots. Various responses to the number of generated shoots were observed when regeneration abilities of different chicory cultivars were examined. Among root and “Witloof” cultivars, ‘Melci’ and ‘Hera’ belong to the root cultivars and exhibited higher shoot regeneration ability. Using the optimized regeneration method, genetic transformation of ‘Melci’ with Agrobacterium tumefaciens strain C58C1 RifR (pGV2260) (pTJK136) was successfully carried out. Histochemical GUS assay, polymerase chain reaction (PCR) and reverse transcription-polymerase chain reaction (RT-PCR) analysis of putative transformed plants confirmed successful integration of the T-DNA into the chicory genome. Expression of the neomycine phosphotransferase (NPTII) in the regenerated plants was also shown by well-developed roots on root inducing medium containing 100 mg l-1 kanamycin. This simple, efficient and reproducible protocol could be useful for inducing somaclonal variation and genetic modification of root chicory cultivars to broaden genetic variation and transferring of important genes