Substomatal ion-adsorbent bodies are reported on here for the first time. A brief survey of the plant kingdom suggests that the structures are mainly restricted to the Commelinaceae and Filicales although analogous structures way occur in other plant groups. Microscopical studies indicate that the bodies have a solid external aspect, a hollow lumen, and are situated extracellularly. In Polypodium, the bodies are narrowly attached to the lower periclinal walls in the polar regions of the guard cell complex, whilst in Tradescantia they are located in the intercellular space between the poles of the complex and the adjacent subsidiary cells. In both genera, the body is covered by the endocuticle which can be distended into a substomatal sac by the body pressing against it. The endocuticle, in the immediate vicinity of the bodies, is modified into a series of hollow trabeculae which are considered to be important apoplastic pathways. The bodies are formed at an early stage of stomatal ontogeny from the migration of outer elements of the lower periclinal wall of the guard-cell mother-cell to both poles of the eventual guard cell complex. The walls of the body are believed to be highly pectinaceous and capable of adsorbing a wide variety of ions non-selectively. Preliminary X-ray microanalyses suggest that the bodies may be involved in potassium fluxes associated with stomatal movements. Ultrastructural studies of immature Polypodium guard cells resulted in the erection of a hypothetical model for stoma formation. Ontogenetic studies revealed a major anomaly in existing stomatal classifications which is rectified in a proposed new classification of stomatal types which is explicit a.t both ontogenetic and morphological levels. Previously unrecorded ontogenetic and morphological stomatal types are reported from Polypodium.

A review of phytosociological techniques, including the Zurich-Montipellier (Z-M) system. Association analysis and simple ordination is made, using a standard data set. The complementary nature of the results obtained is noted. As classification erects divisions and boundaries in a data set, and the prime method used in the analysis of heath vegetation is classificatory, evidence for the prescence of 'real' vegetation boundaries is presented, together with a discussion of their nature and function. Heathland vegetation in Britain has been classified using the Z-M phytosociological system, and four main types have been distinguished, which are equated with alliances at present in use in Europe. These are; Erica cinerea Heath: Ulicion nanae (Duvigud, 1944)em. Van den Bergen. Calluna vulgaris Heath: Calluno-Genisition pilosae (Duvignd, 1944) Vaccinium myrtillus Heath: Llyrtillion bore ale (Bocher, 1943). Erica tetralix Heath: Ericion tetralicis (Schuick, 1933).The first three are typical of Dry heaths, whereas the fourth indicates wet heath. A phytogeographical analysis of the Heath vegetation gives further validity to these four types, indicating the dependence of the three dry 'Heaths' on species which are geographically restricted. Use of the Z-M system and some other techniques to indicate the nature of recent vegetation in areas now urbanised or agriculturalised is made, using South Gloucestershire as such an area.

Vegetation is abundant in rivers, and has a significant influence on their hydraulic, geomorphological, and ecological functioning. However, past modelling of the influence of vegetation has generally neglected the complexity of natural plants. This thesis develops a novel numerical representation of flow through and around floodplain and riparian vegetation, focusing on flow-vegetation interactions at the plant-scale. The plant volumetric canopy morphology, which comprises the distribution of vegetal elements over the three-dimensional plant structure, is accurately captured at the millimetre scale spatial resolution using Terrestrial Laser Scanning (TLS), and incorporated into a Computational Fluid Dynamics (CFD) model used to predict flow. Numerical modelling, with vegetation conceptualised as a porous blockage, is used to improve the process-understanding of flow-vegetation interactions. Model predictions are validated against flume experiments, with plant motion dynamics investigated, and analysis extended to consider turbulent flow structures and the plant drag response. Results demonstrate the spatially heterogeneous velocity fields associated with plant volumetric canopy morphology. The presence of leaves, in addition to the posture and aspect of the plant, significantly modifies flow field dynamics. New insights into flow-vegetation interactions include the control of plant porosity, influencing ‘bleed-flow’ through the plant body. As the porosity of the plant reduces, and bleed-flow is prevented, the volume of flow acceleration increases by up to ~150%, with more sub-canopy flow diverted beneath the impermeable plant blockage. Species-dependent drag coefficients are quantified; these are shown to be dynamic as the plant reconfigures, differing from the commonly assigned value of unity, and for the species’ investigated in this thesis range between 0.95 and 2.92. The newly quantified drag coefficients are used to re-evaluate vegetative flow resistance, and the physically-determined Manning’s n values calculated are highly applicable to conveyance estimators and industry standard hydraulic models used in the management of the river corridor.

Extensive critical review of literature and stakeholder interrogation provided key research questions and paradigms. They are explained in the introductory chapters. Approaches to the understanding and assessment of woods and of hedgerows (as linear ‘woodlands’) were developed and tested through intensive and extensive field-based case studies. This research investigated and critically assessed the role and value of using botanical indicators as historic markers in wooded landscapes that comprise woodlands and hedgerows. These are linked by social history and ecology. In both habitat types, there have been recent attempts to determine their age and origins based on current floras. Ancient woodlands (i.e. present pre-1600) are determined by reference to regional ancient woodland indicator species (AWI) lists. Hedgerows have been dated by counting the number of woody species in sections (the Hooper Rule) to provide an estimate of hedgerow age. In this study, both the derivation of ancient woodland indicator species and the dating of hedgerows using the 'Hooper Rule' were questioned. In particular, the survey methods applied in these situations were critically analysed. For woodlands, there has been only limited emphasis on recording the local variations in flora within woodland. The woody species counting for hedgerows took little account of the species involved. Stakeholder opinion was canvassed using a series of four woodland workshops where the role of AWI was discussed. This generated questions the outcomes of which agreed with this research that new methods of data collection and interpretation were needed. Furthermore, the current patterns of the use of ancient woodland indicator species at regional or county level were considered and assessed. The need for a new approach to surveying woodlands and hedgerows to collect data relevant to historic interpretation was addressed. Appropriate methodologies were proposed and tested. A novel approach to interpretation was developed that considered the nature of a species used as an historic marker: where it was, how abundant it was and if there were any other associated species in combination. This intelligent interrogation process is a radical departure from current approaches to using only the presence of botanical species as historic markers. The overall conclusion of this research is that botanical species are valuable and powerful historic markers if their presence is considered carefully and intelligently based on adequately detailed surveys. This original approach has added to scientific knowledge and the understanding of botanical species as historic markers. New practitioner and researcher toolkits were developed and tested, and novel approaches to the evaluation of woods and hedgerows using cross-disciplinary methods were proposed.

Potassium-deficient plants commonly have much higher respiration rates than their high-potassium controls. In view of the evidence that respiratory intensity is largely controlled by the internal level of phosphate acceptors like adenosine diphosphate, an attempt was made to determine whether potassium deficiency does in fact affect the free nucleotide contents of plants in any such manner as would explain the respiratory effect along these lines. Serious difficulties of technique •were encountered in estimating the nucleotides, and these have not been completely satisfactorily overcome. Certain widely used techniques were found wanting in various respects. Precipitation methods involving barium and zinc wore tested on both authentic adenine nucleotides and leaf preparations. These were abandoned, owing principally to variable recoveries of authentic nucleotides and to the presence of large quantities of precipitable impurities in leaf extracts. Precipitation with ethanol, a commonly used method for removing such interfering substances, was found to cause considerable losses of the nucleotides. Selective adsorption of nucleotides by various charcoals was found to induce an anomalous behaviour of AMP as regards its elution position when chromatographed on Dowex-1-chloride. This effect, whose nature is unknown, was a property of all the charcoals tested. Water extraction, following a short pre-immersion in boiling ethanol, was adopted for most of the analyses, but perchloric acid was sometimes used as the extractant; this acid -ma shown to release considerable quantities of bound nucleotides. Analyses of the free nucleotides in leaves of high- and low-potassium barley by chromatography on Dowex-1-Chloride and formate columns furnished no evidence of significant changes therein as a result of potassium deficiency. It is tentatively concluded that the enhanced respiration of potassium deficient barley must be primarily due to factors other than the levels of detectable free nucleotides.

Calcutta Botanic Garden was founded in 1786 to acclimatise economic plants, but it quickly became the main institutional base for scientific botany in colonial India. However, it had to make a new start in 1833 after the Garden superintendent, Nathaniel Wallich, distributed its herbarium to botanists in Europe. The thesis shows how the revival of the scientific project to investigate and catalogue the south Asian flora was the main priority for Wallich’s successors, but depended on successful negotiation with the government. The central theme of the thesis is the tension between scientists, intent on their research, and sponsors, who need to demonstrate practical outcomes. It breaks new ground by focussing on how these issues were debated and resolved within a particular colonial scientific institution. It argues that the Garden was able to attract the resources it needed for its scientific work by responding appropriately to government pressures: although its achievements in economic botany were limited, it successfully highlighted them, regularly citing the introduction of tea and cinchona; it reinforced its case by managing its site in ways that reassured the government. The thesis also adds to our understanding of centre-periphery relationships. It argues that the Garden’s role as an important nodal point in the global botanic network was key to achieving its objectives. It shows how the Garden was strengthened by its mutually supportive relationship with Kew Gardens, based on the close bonds that botanists formed with each other. The thesis concludes by showing how, despite the Garden’s achievements, the government gradually lost faith in the ability of botany to contribute to economic progress in India; in the twentieth century it increasingly turned to more specialist disciplines and institutions. The thesis therefore suggests that further studies of scientific institutions would enhance our understanding of how science continued to support and validate imperial rule.

Manganese (Mn) is an essential heavy metal micronutrient in plant growth and development, but becomes toxic when present in excess, with potential agricultural yield losses under both nutritional extremes. Membrane transporters play a key role in enabling plant growth under Mn deficiency and toxicity. This study directly compares the roles of members from different ubiquitous transporter families in Mn homeostasis: Group 8/9 members of the Metal Tolerance Proteins (MTPs), At MTP8–MTP11; At ECA3, a P2A-type ATPase; and Natural Resistance Associated Macrophage Proteins, At NRAMP1 and At NRAMP2. MTP8 plays a significant role in Mn detoxification at the tonoplast, conferring Mn hypertolerance when overexpressed; it is also the only Group 8/9-MTP involved in alleviating Mn/iron antagonism. A role in Mn detoxification is also assigned to MTP10, identified through comparison of novel double and triple mtp knockout mutants. Expression in yeast provides evidence that these proteins can transport Mn, with greater Mn tolerance conferred by MTP8 and MTP10 than MTP11. ECA3 is also shown to play a minor role in conferring tolerance to Mn toxicity in planta, but this is only apparent when MTP11 is non-functional; its major role appears to lie in alleviating Mn deficiency. This thesis also clarifies the disputed subcellular localisations of ECA3 and MTP11, demonstrating targeting to the trans- and cis-Golgi, respectively. Additionally, NRAMP2 is shown to alleviate Mn deficiency at the cis-Golgi. Characterisation of double and triple mutants has begun to clarify the relative contribution of NRAMP1, NRAMP2 and ECA3 in Mn deficiency. The use of mutants in this study has also highlighted the antagonistic effects of calcium in Mn homeostasis. Overall, this study provides a more comprehensive understanding of how key transporters function together in Mn homeostasis, identifying targets that may be useful for crop improvement.

Phylogenetics focuses on learning about evolutionary relationships between species. These relationships can be represented by phylogenetic trees, where similar species are grouped together as sharing a recent common ancestor. The common ancestor of all the species of the tree is the root of the tree. The root is fundamental to the biological interpretation of the tree, providing a critical reference point for polarising ancestor-descendant relation- ships and determining the order in which key traits evolved along the tree (Embley and Martin, 2006). Despite its importance, most models of sequence evolution are unable to infer the root of a phylogenetic tree. They are based on homogeneous continuous time Markov processes (CTMPs) that are assumed to be stationary and time-reversible, with the mathematical consequence that the likelihood of a tree does not depend on where it is rooted. As a result, the root of the tree cannot be inferred as part of the analysis. Other methods which are generally used to root evolutionary trees can be problematic. For example, the outgroup rooting method is susceptible to a long-branch attraction arte- fact. Paralogue rooting requires pairs of paralogous genes which underwent an ancient gene duplication event to be present in all species being analysed, and the number of such genes is limited. In this thesis we explore an alternative model-based approach, adopting a substitution model in which changing the root position changes the likelihood of the tree. We explore the e ect of relaxing reversibility and stationarity assumptions and allowing the position of the root to be another unknown quantity in the model. We propose two hierarchical non-reversible models which are centred on a reversible model but perturbed to allow non- reversibility. The models di er in the degree of structure imposed on the perturbations. We also explore non-stationary models, and the combination of relaxing both the reversibility and the stationarity assumptions. The analysis is performed in the Bayesian framework using Markov chain Monte Carlo methods. We illustrate the performance of the models in analyses of simulated datasets using two types of topological priors. We also investigate the e ect of di erent topologies and branch lengths on the inference. Our results illustrate the usefulness of modelling non- reversibility and non-stationarity for root inference, and also demonstrate the sensitivity of the analysis to topological priors. We then apply the models to real biological datasets, the radiation of polyploid yeasts and the radiation of primates, for which there is a robust biological opinion about the root position. Finally we apply the models to an open question in biology: rooting the ribosomal tree of life.

Crassulacean acid metabolism (CAM) is a metabolic adaptation of photosynthesis that is optimised via strict temporal regulation of its biochemistry by the circadian oscillator. CAM plants achieve high water use efficiency and thus thrive in seasonally dry regions unsuitable for C3 food crops such as rice or wheat. Climate change and the associated challenges of global food and energy security, mean that CAM research is currently of urgent and pressing need, as CAM may reveal methods for the generation of more water use efficient crops. The efficiency of CAM is optimised by the circadian clock, through the regulation of PHOSPHOENOLPYRUVATE CARBOXYLASE KINASE (PPCK) expression and nocturnal CO2 fixation, but the signalling pathway between the central clock and CAM has yet to be elucidated. Whole genome sequencing and detailed RNA-seq datasets for C3 and CAM leaves of the model CAM species Kalanchoë fedtschenkoi have enabled the discovery of novel genes that could function to link CAM to the circadian clock. Three CAM-induced and clock-controlled transcription factor (TF) genes were identified from the RNA-seq datasets: MYB-LIKE 439 (KfMYB439), CAM-INDUCED BZIP1 (KfCIB1) and CYCLING DOF FACTOR2 (KfCDF2). Both over-expressor and RNAi knockdown transgenic lines of K. fedtschenkoi were generated for each TF, facilitating the further elucidation of their biological functions. Over 200 transgenic lines were screened for altered expression levels, and changes to the dawn and dusk levels of key CAM metabolites: malate and starch. Four transgenic lines for each TF, two over-expressor and two RNAi lines, were used for detailed phenotypic analysis of CAM-associated traits. Transgenic perturbation of any one of the three TFs caused small but widespread changes to the transcript levels of core clock- and CAM-associated genes. KfMYB439 is a REVEILLE family TF related to the clock gene CIRCADIAN CLOCK ASSOCIATED1 (CCA1). Data revealed that KfMYB439 functioned close to the core circadian oscillator. Mis-regulation of KfMYB439 led to the perturbation of efficient dark CO2 fixation, reduced levels of starch and malate, and reduced productivity during drought. KfCIB1, was found to feed back to influence the core circadian clock as well as regulating CAM. In constant light conditions, KfCIB1 mis-expression led to perturbed timing of KfCCA1 and TIMING OF CAB1 (KfTOC1) . KfCIB1 mis-regulation also impacted on stomatal control. At dusk and dawn, large and rapid changes in stomatal conductance resulted in spikes of CO2. Mis-expression also resulted in small improvements in productivity in water-limited environments. KfCDF2 was found to function not only in the clock control of CAM, but also in the photoperiodic control of flowering time. In terms of CAM and the clock, KfCDF2 mis-expression caused changes to CCA1, TOC1 and PPCK expression, and arrhythmic CO2 fixation in constant conditions. It also impacted on water retention during drought, with both over-expressor and RNAi lines displaying higher succulence than the wild type lines after 90 days of drought. KfCDF2 over-expression in both K. fedtschenkoi and K. laxiflora caused constitutive flowering in long days, whereas wild type plants never flowered. Q-RT-PCR analysis of flowering pathway genes revealed that KfCDF2 over-expression impacted on transcript levels for CONSTANS (CO) and FLOWERING LOCUS T (FT); key proteins in the day-length dependent induction of flowering. Results suggested that all three TFs likely function in the circadian optimisation of CAM, although whether or not the often small effects were direct or indirect will require further work. Future Chromatin Immunoprecipitation and sequencing experiments will reveal the target genes regulated by these TFs, and the identification of other novel CAM-induced genes from the RNA-seq data, will allow the pathway between the circadian clock and CAM to be elucidated in much greater detail.

The tobacco NtmybAS1 gene encodes a novel anther-specific member of the Myb family of transcription factors. Overexpression of NtmybAS1 driven by the CaMV35S promoter in tobacco resulted in dramatic alterations in plant architecture leading to a semi-dominant dwarf phenotype. Northern blot analysis demonstrated a direct relationship between the severity of dwarf phenotype and the level of NtmybAS1 transcripts, which clearly indicated that the NtmybAS1-induced dwarfing is gene dosage dependent. Analysis of cell morphology demonstrated that a severe reduction of cell elongation in hypocotyls was the major cause of NtmybAS1-induced dwarfing. In contrast, a dramatic increase in cell elongation and pronounced cell division was detected in palisade and mesophyll cells. Despite changes in cell morphology, the overall body organisation of NtmybAS1 plants remained unaffected. Furthermore, phenotypic alterations in NtmybAS1 overexpressing plants were not normalised by application of phytohormones or by grafting, indicating the involvement of non-hormonal factors and the cell autonomy of dwarfing. Transient expression analyses of the N-terminal Myb domain derivatives fused to sGFP revealed the presence of a nuclear localisation signal, which efficiently targeted sGFP to the pollen nucleus. Analysis of the cellular protein profiles of NtmybAS1 plants by one and two-dimensional gel electrophoresis revealed two proteins that were highly induced in NtmybAS1 plants. Tryptic peptides derived from these proteins by reversed-phase HPLC were sequenced and showed an exact match with two pathogenesis-related (PR) proteins, tobacco chitinase P (PR-P) and PR-1a.