Genetic engineering of the forage legume lotus corniculatus using Agrobacterium: mediated transformation systems

Gene transfer vectors based on the Agrobacterium tumefaciens Ti plasmid were used to develop a successful disarmed Agrobacterium tumefaciens-mediated transformation method for Lotus comiculatus. A binary vector construct, pJIT73, was used during the development of the Agrobacterium tumefaciens transformation system due to its selectable (Aph IV, nos- neo) and scorable markers. The effects of the antibiotics geneticin (G-418) and hygromycin B were studied. Use of kill curves and selection delay experiments allowed potentially suitable selection pressure parameters to be proposed. Using such selection during transformation experiments led to further optimisation of this stage of transformation. The influence of plant hormones on the regeneration of Lotus comiculatus explants was investigated and a modification of an established protocol using leaf explants was introduced as an attempt to reduce the overall time of regeneration. Various explants were used but leaf pieces were chosen as the most suitable explant on which to focus research. So, through alteration of various stages, including length of cocultivation and subsequent decontamination within the transformation process, a successful method was developed. Experiments indicated the optimum Agrobacterium tumefaciens strain to be used with Lotus comiculatus was the disarmed Ach5 type, LBA4404(pAL4404). Transgenic Lotus comiculatus plants were produced which expressed the scorable marker β-Glucuronidase gene (GUS) and the selectable marker for hygromycin B resistance, AphIV. Gene transfer was confirmed by Southern blotting. The new Agrobacterium tumefaciens-mediated vector system was used to introduce the cowpea trypsin inhibitor gene (CpTi) into Lotus comiculatus. However, although there was evidence for transformed callus development, no shoots were induced. By the use of previously established Agrobacterium rhizogenes-mediated system, an attempt was made to introduce the pea lectin gene (psl) into Lotus corniculatus. Hairy root regenerants were produced but genetic transfer was unconfirmed and attempted investigation of the plant - Rhizobium symbiosis involving Lotus corniculatus was not fulfilled

Vol. 14, No. 2

Contents: New Developments in Public Sector Arbitration—The New Trilogy and Much Ado About Nothing, by Gregory J. Malovance, Margaret R. Gibbs Recent Developments, by the Student Editorial Board Further References, compiled by Margaret A. Chaplanhttps://scholarship.kentlaw.iit.edu/iperr/1027/thumbnail.jp

Vol. 14, No. 2

Contents: New Developments in Public Sector Arbitration—The New Trilogy and Much Ado About Nothing, by Gregory J. Malovance, Margaret R. Gibbs Recent Developments, by the Student Editorial Board Further References, compiled by Margaret A. Chaplanhttps://scholarship.kentlaw.iit.edu/iperr/1027/thumbnail.jp

The interactions between macrophytes and sediments in urban river systems.

PhDMany urban rivers receive significant inputs of metal-contaminated sediments from their catchments. Their restoration has the potential to increase the deposition and accumulation of these sediments from greater sediment supply and increased channel hydraulic complexity, creating a store of metals which could have negative impacts upon ecosystems and human health. Macrophytes often establish in restored channels and have the potential to stabilise these sediments and uptake metals through processes of phytoremediation, thus reducing the risk of the accumulated sediments becoming a source of metals. This thesis investigates the effects of river restoration upon sedimentation patterns and the interactions between macrophytes and sediments in terms of sediment trapping, stabilisation and metal uptake within urban river systems. At a reach scale, greater finer sediment deposition and the accumulation of sediment around in-channel vegetation was found within restored stretches of tributaries of the River Thames London, reflecting sediment availability and hydraulic conditions. These sediments were important in terms of greater metal storage within stretches, and along with gravels showed particularly high metal concentrations. Interactions between macrophytes, sediment and flow were investigated within the urban-influenced River Blackwater, Surrey. At the stand scale, the common emergent Sparganium erectum was found to significantly reduce flow velocities, accumulate fine sediments and retain them over winter. Research on individual plants revealed that, although three common emergent macrophytes (Sparganium erectum, Typha latifolia and Phalaris arundinacea) did not significantly phytoremediate metal contaminated sediments through metal uptake or bioconcentration, the reinforcement and stabilisation of these accumulated sediments (particularly by Sparganium erectum and Typha latifolia) and the creation of anoxic sediment conditions which strongly bind metals, were important in reducing the risk of metal mobilisation from the sediments. These macrophyte sediment interactions illustrate the great potential of using emergent macrophytes in the restoration and management of urban rivers with metal contaminated sediments

Notch ligation by Delta1 inhibits peripheral immune responses to transplantation antigens by a CD8⁺ cell–dependent mechanism

Notch signaling plays a fundamental role in determining the outcome of differentiation processes in many tissues. Notch signaling has been implicated in T versus B cell lineage commitment, thymic differentiation, and bone marrow hematopoietic precursor renewal and differentiation. Notch receptors and their ligands are also expressed on the surface of mature lymphocytes and APCs, but the effects of Notch signaling in the peripheral immune system remain poorly defined. The aim of the studies reported here was to investigate the effects of signaling through the Notch receptor using a ligand of the Delta-like family. We show that Notch ligation in the mature immune system markedly decreases responses to transplantation antigens. Constitutive expression of Delta-like 1 on alloantigen-bearing cells renders them nonimmunogenic and able to induce specific unresponsiveness to a challenge with the same alloantigen, even in the form of a cardiac allograft. These effects could be reversed by depletion of CD8⁺ cells at the time of transplantation. Ligation of Notch on splenic CD8⁺ cells results in a dramatic decrease in IFN-γ with a concomitant enhancement of IL-10 production, suggesting that Notch signaling can alter the differentiation potential of CD8⁺ cells. These data implicate Notch signaling in regulation of peripheral immunity and suggest a novel approach for manipulating deleterious immune responses