The cellular basis of the elongation response in submerged deep-water rice

The cellular basis of internode elongation was studied in intact deep-water rice plants (Oryza sativa L. cv. Habiganj Aman II) and in isolated stem sections. In intact plants, growth was stimulated by submergence in water and by ethylene treatment. In isolated sections, growth was enhanced by submergence, by ethylene, and by exposure of the tissue to an atmosphere of 3% O₂, 91% N₂ and 6% CO₂ or 3% O₂, 91% N₂, 6% CO₂ and 1 l l^-1 C₂H₄ (by vol.). Under all these conditions, growth was localized in the intercalary meristem at the bases of the internodes. Autoradiography of [³H]thymidine-labeled tissue showed activation of cell division and longitudinal expansion of the intercalary meristem. Increased production of new cells and their subsequent elongation thus form the basis for the growth response to submergence and ethylene treatment in deep-water rice plants.     

Amylase activity and growth in internodes of deepwater rice

Isoelectrofocusing, product analysis, thermal denaturation studies and affinity chromatography on cycloheptaamylose-Sephadex were used to identify the amylolytic enzymes in internodes of deepwater rice (Oryza sativa L.). Amylolytic activity in internodes of deepwater rice consists of -amylase (sometimes separated into two isoforms) and of -amylase. During submergence of whole plants, -amylase activity increases in young, growing internodes, but -amylase activity declines. Although non-growing, mature internodes contain higher levels of -amylase than do the elongating younger internodes, the effect of submergence on amylase activities in both tissues follows the same trend. Submergence, gibberellic acid (GA₃) and ethylene all promote -amylase activity in growing and non-growing internodes of excised deepwater-rice stem sections. Inhibitor studies showed that submergence and ethylene promote -amylase activity in the absence of endogenous gibberellin (GA), and GA₃ enhances -amylase activity when ethylene action is inhibited. Therefore, ethylene and GA appear to increase -amylase activity independently of each other. Enhanced -amylase activities are probably responsible for the mobilization of carbohydrates which are needed to support internode elongation during submergence of deepwater rice.Abbreviations CHA cycloheptaamylose - GA₃ gibberellic acid - NBD 2,5-norbornadiene - TCY tetcyclacis     

The effect of submergence,ethylene and gibberellin on polyamines and their biosynthetic enzymes in deepwater-rice internodes

Submergence and treatment with ethylene or gibberellic acid (GA₃) stimulates rapid growth in internodes of deepwater rice (Oryza sativa L. cv. Habiganj Aman II). This growth is based on greatly enhanced rate of cell-division activity in the intercalary meristem (IM) and on increased cell elongation. We chose polyamine biosynthesis as a biochemical marker for cell-division activity in the IM of rice stems. Upon submergence of the plant, the activity of S-adenosylmethionine decarboxylase (SAMDC; EC 4.1.1.50) in the IM increased six- to tenfold within 8 h; thereafter, SAMDC activity declined. Arginine decarboxylase (ADC; EC 4.1.1.19) showed a similar but less pronounced increase in activity. The activity of ornithine decarboxylase (ODC; EC 4.1.1.17) in the IM was not affected by submergence. The levels of putrescine and spermidine also rose in the IM of submerged, whole plants while the concentration of spermine remained low. The increase in SAMDC activity was localized in the IM while the activity of ADC rose both in the node and the IM above it. The node also contained low levels of ODC activity which increased slightly following submergence. Increased activities of polyamine-synthesizing enzymes in the nodal region of submerged plants probably resulted from the promotion of adventitious root formation in the node. Treatment of excised rice-stem sections with ethylene or GA₃ enhanced the activities of SAMDC and ADC in the IM and inhibited the decline in the levels of putrescine and spermidine. We conclude that SAMDC and perhaps also ADC may serve as biochemical markers for the enhancement of cell-division activity in the IM of deepwater rice.Abbreviations ADC arginine decarboxylase - GA gibberellin - IM intercalary meristem - ODC ornithine decarboxylase - SAM S-adenosylmethionine - SAMDC SAM decarboxylase     

Low levels of oxygen promote internodal elongation in floating rice independently of enhanced ethylene production

Submergence induces rapid elongation of internodes in floating rice(Oryza sativa L. cv. Habiganj Aman II). The initial signalfor such internodal elongation has been considered to be the reduced partialpressure of oxygen in submerged internodal cavities, which promotes theelongation of internodes through the enhancement of ethylene synthesis. Weexamined the relationship between low oxygen pressure and ethylene production inthe rapid elongation of floating rice internodes using ethylene biosynthesisinhibitors, aminooxyacetic acid (AOA) and CoCl₂. When floating ricestem segments were incubated in an atmosphere of low O₂, internodalelongation accelerated and ethylene production increased. However, in stemsegments treated with AOA or CoCl₂, low O₂ stillstimulated the elongation of internodes although the ethylene production by theinternodes was less than by those in control stem segments where internodalelongation was not promoted. These results indicate that low O₂ iscapable of causing rapid elongation of internodes of floating rice independentlyof enhanced production of ethylene. In addition to low O₂,submergence, ethylene and gibberellic acid each enhanced the production ofethylene by internodal tissues, suggesting that enhanced ethylene production isa common phenomenon accompanied by the acceleration of internodal elongation infloating rice.     

Internodal elongation and orientation of cellulose microfibrils and microtubules in deepwater rice

Excised stem sections of deepwater rice (Oryza sativa L.) containing the highest internode were used to study the induction of rapid internodal elongation by gibberellin (GA). It has been shown before that this growth response is based on enhanced cell division in the intercalary meristem and on increased cell elongation. In both GA-treated and control stem sections, the basal 5-mm region of the highest internode grows at the fastest rate. During 24 h of GA treatment, the internodal elongation zone expands from 15 to 35 mm. Gibberellin does not promote elongation of internodes from which the intercalary meristem has been excised. The orientation of cellulose microfibrils (CMFs) is a determining factor in cell growth. Elongation is favored when CMFs are oriented transversely to the direction of growth while elongation is limited when CMFs are oriented in the oblique or longitudinal direction. The orientation of CMFs in parenchymal cells of GA-treated and control internodes is transverse throughout the internode, indicating that CMFs do not restrict elongation of these cells. Changes in CMF orientation were observed in epidermal cells, however. In the basal 5-mm zone of the internode, which includes the intercalary meristem, CMFs of the epidermal cell walls are transversely oriented in both GA-treated and control stem sections. In slowly growing control internodes, CMF orientation changes to the oblique as cells are displaced from this basal 5-mm zone to the region above it. In GA-treated rapidly growing internodes, the reorientation of CMFs from the transverse to the oblique is more gradual and extends over the 35-mm length of the elongation zone. The CMFs of older epidermal cells are obliquely oriented in control and GA-treated internodes. The orientation of the CMFs parallels that of the cortical microtubules. This is consistent with the hypothesis that cortical microtubules determine the direction of CMF deposition. We conclude that GA acts on cells that have transversely oriented CMFs but does not promote growth of cells whose CMFs are already obliquely oriented at the start of GA treatment.     

Cell-wall synthesis and elongation growth in hypocotyls of Helianthus annuus L.

The relationship between growth and increase in cell-wall material (wall synthesis) was investigated in hypocotyls of sunflower seedlings (Helianthus annuus L.) that were either grown in the dark or irradiated with continuous white light (WL). The peripheral three to four cell layers comprised 30–50% of the entire wall material of the hypocotyl. The increase in wall material during growth in the dark and WL, respectively, was larger in the inner tissues than in the peripheral cell layers. The wall mass per length decreased continuously, indicating that wall thinning occurs during growth of the hypocotyl. When dark-grown seedlings were transfered to WL, a 70% inhibition of growth was observed, but the increase in wall mass was unaffected. Likewise, the composition of the cell walls (cellulose, hemicellulose, pectic substances) was not affected by WL irradiation. Upon transfer of dark-grown seedlings into WL a drastic increase in wall thickness and a concomitant decrease in cell-wall plasticity was measured. The results indicate that cell-wall synthesis and cell elongation are independent processes and that, as a result, WL irradiation of etiolated hypocotyls leads to a thickening and mechanical stiffening of the cell walls.     

Cell-lineage patterns in the shoot apical meristem of the germinating maize embryo

A fate map for the shoot apical meristem of Zea mays L. at the time of germination was constructed by examining somatic sectors (clones) induced by -rays. The shoot apical meristem produced stem, leaves, and reproductive structures above leaf 6 after germination and the analysis here concerns their formation. On 160 adult plants which had produced 17 or 18 leaves, 277 anthocyanin-deficient sectors were scored for size and position. Sectors found on the ear shoot or in the tassel most often extended into the vegetative part of the plant. Sectors ranged from one to six internodes in length and some sectors of more than one internode were observed at all positions on the plant. Single-internode sectors predominated in the basal internodes (7,8,9) while longer sectors were common in the middle and upper internodes. The apparent number of cells which gave rise to a particular internode was variable and sectors were not restricted to the lineage unit: a leaf, the internode below it, and the axillary bud and prophyll at the base of the internode. These observations established two major features of meristem activity: 1) at the time of germination the developmental fate of any cell or group of cells was not fixed, and 2) at the time of germination cells at the same location in a meristem could produce greatly different amounts of tissue in the adult plant. Consequently, the developmental fate of specific cells in the germinating meristem could only be assigned in a general way.Abbreviations ACN apparent cell number - LI, LII, LI-LII sectors restricted to the epidermis, the subepidermis, or encompassing epidermis and subepidermis - PCN progenitor cell     

Spatial distribution of growth rates and of epidermal cell lengths in the elongation zone during leaf development in Lolium perenne L.

Relative elemental growth rates (REGR) and lengths of epidermal cells along the elongation zone of Lolium perenne L. leaves were determined at four developmental stages ranging from shortly after emergence of the leaf tip to shortly before cessation of leaf growth. Plants were grown at constant light and temperature. At all developmental stages the length of epidermal cells in the elongation zone of both the blade and sheath increased from 12 m at the leaf base to about 550 m at the distal end of the elongation zone, whereas the length of epidermal cells within the joint region only increased from 12 to 40 m. Throughout the developmental stages elongation was confined to the basal 20 to 30 mm of the leaf with maximum REGR occurring near the center of the elongation zone. Leaf elongation rate (LER) and the spatial distributions of REGR and epidermal cell lengths were steady to a first approximation between emergence of the leaf tip and transition from blade to sheath growth. Elongation of epidermal cells in the sheath started immediately after the onset of elongation of the most proximal blade epidermal cells. During transition from blade to sheath growth the length of the blade and sheath portion of the elongation zone decreased and increased, respectively, with the total length of the elongation zone and the spatial distribution of REGR staying near constant, with exception of the joint region which elongated little during displacement through the elongation zone. Leaf elongation rate decreased rapidly during the phase when only the sheath was growing. This was associated with decreasing REGR and only a small decrease in the length of the elongation zone. Data on the spatial distributions of growth rates and of epidermal cell lengths during blade elongation were used to derive the temporal pattern of epidermal cell elongation. These data demonstrate that the elongation rate of an epidermal cell increased for days and that cessation of epidermal cell elongation was an abrupt event with cell elongation rate declining from maximum to zero within less than 10 h.Abbreviations LER leaf elongation rate - REGR relative elemental growth rates     

Persistent effects of changes in phytochrome status on internode growth in light-grown mustard: Occurrence,kinetics and locus of perception

Extension growth of the first internode in fully de-etiolated mustard (Sinapis alba L.) seedlings (11–12.5 d old) is under the control of both the current phytochrome photoequilibrium (Pfr/P, ratio of the far-red-absorbing form of phytochrome to total phytochrome) and that established by short (<12 h) pretreatments. Plants were pretreated with either light pulses providing different calculated Pfr/P followed by dark incubations of different durations (a), or with a 12-h period of white light establishing different Pfr/P (b). After the pretreatments, the plants received either light pulses providing different Pfr/P, followed by dark incubations (c), or continuous white light with or without addtional far-red light (d). Thus, four experimental approaches were followed: (a)(c); (a)(d); (b)(c) and (b)(d). Extension growth during the second period (c or d) was not only affected by the current phytochrome status, but also by that established during the pretreatment period (a or b). The results show the existence of a long-term promotion of stem growth which persists after the end of the low Pfr/P pretreatment. This effect is different from the previously reported rapid effect of far-red light added to background white light as follows: (i) the duration of low Pfr/P required to effect a full response is longer (2.5 h); (ii) the duration of the promotion after returning to high Pfr/P is longer (approx. 24 h) and (iii) the locus of perception is mainly in the leaves, rather than the growing internode.Abbreviations FR far-red light - PAR photosynthetically active radiation - Pfr/P ratio between the FR-absorbing form and total phytochrome - R red light - WL white light     

A kinematic analysis of tepal growth in Lilium longiflorum

Time-lapse marking experiments indicate that the growth of tepals in Lilium longiforum Thunb. from 3.7 mm to maturity is triphasic. Phase I (tepal lengths 3.7–10 mm) is characterized by spatial and temporal variation in growth rate and, in the epidermis, a random distribution of mitoses with an acropetal increase in cell area. During phase II (10–90 mm) cell elongation and (later) cell division is restricted largely to basal regions. Cell division ceases when tepals are less than one-third of their mature length of 155 mm. Phase III (90–155 mm) is characterized by the gradual transition from basal to apical growth, and a modification of epidermal cell shape. A sharp peak in growth at the extreme tip of the tepal coincides with anthesis.Abbreviations LRGR local relative growth rate - RER relative elemental rate of growth     

Phytochrome inhibits the effectiveness of gibberellins to induce cell elongation in rice

Red light controls cell elongation in seedlings of rice (Oryza sativa L.) in a far-red-reversible manner (Nick and Furuya, 1993, Plant Growth Regul. 12, 195–206). The role of gibberellins and microtubules in the transduction of this response was investigated in the rice cultivars Nihon Masari (japonica type) and Kasarath (indica type). The dose dependence of mesocotyl elongation on applied gibberellic acid (GA₃) was shifted by red light, and this shift was reversed by far-red light. In contrast, coleoptile elongation was found to be independent of exogenous GA₃. Nevertheless, it was inhibited by red light, and this inhibition was reversed by far-red light. The content of the active gibberellin species GA₁ and GA₄ was estimated by radio-immunoassay. In the mesocotyl, the gibberellin content per cell was found to increase after irradiation with red light, and this increase was far-red reversible. Conversely, the cellular gibberellin content in japonica-type coleoptiles did not exhibit any significant light response. Microtubules reoriented from transverse to longitudinal arrays in response to red light and this reorientation could be reversed by subsequent far-red light in both the coleoptile and the mesocotyl. This movement was accompanied by changes in cell-wall birefringence, indicating parallel reorientations of cellulose deposition. The data indicate that phytochrome regulates the sensitivity of the tissue towards gibberellins, that gibberellin synthesis is controlled in a negative-feedback loop dependent on gibberellin effectiveness, and that at least two hormone-triggered signal chains are linked to the cytoskeleton in rice.Abbreviations D darkness - FR far-red light - GA₃ gibberellic acid - GC-SIM gas chromatography-selected ion monitoring - R red light This work was supported by a grant of the Human Frontier Science Organization to P.N. Advice and organizational support by Prof. M. Furuya (Hitachi Advanced Research Laboratory, Hatoyama, Japan) and Prof. N. Murofushi (Department of Agricultural Chemistry, University of Tokyo, Japan) is gratefully acknowledged. Seeds of both rice cultivars were kindly provided by Dr. O. Yatou (Institute for Radiation Breeding, Hitachi-Ohmiya, Japan), and the antiGA₁ Me-antiserum for the radio-immunoassays by Dr. I. Yamaguchi (Department of Agricultural Chemistry, University of Tokyo, Japan).     

Regulation of growth in stem sections of deep-water rice

Submergence in water greatly stimulates internodal elongation in excised stem sections of deep-water rice (Oryza sativa L. cv. Habiganj Aman II) and inhibits growth of leaf blades and leaf sheaths. The highest rates of internodal growth have been observed in continuous light. Very little growth occurs in submerged sections kept in darkness or incubated under N₂ in the light. The effect of submergence on the growth of deep-water rice is, at least in part, mediated by C₂H₄, which accumulates in the air spaces of submerged sections. This accumulation results from increased C₂H₄ synthesis in the internodes of submerged sections and reduced diffusion of C₂H₄ from the tissue into the water. Increased C₂H₄ levels accelerate internodal elongation and inhibit the growth of leaves. Compounds capable of changing the rate of C₂H₄ synthesis, namely aminoethoxyvinylglycine, an inhibitor of C₂H₄ synthesis, and 1-aminocyclopropane-1-carboxylic acid, the immediate, precursor of C₂H₄, have opposite effects on growth of internodes and leaves. The enhancement of internodal elongation by C₂H₄ is particularly pronounced in an atmosphere of high CO₂ and low O₂. The increase in C₂H₄ synthesis in internodes of submerged sections is primarily triggered by reduced atmospheric concentrations of O₂. The rate of C₂H₄ evolution by internodes isolated from stem sections and incubated in an atmosphere of low O₂ is up to four times greater than that of isolated internodes incubated in air. In contrast, C₂H₄ evolution from the leaves is reduced under hypoxic conditions. The effect of submergence on growth of stem sections of deep-water rice can be mimicked by exposing non-submerged sections to a gas mixture which is similar to the gaseous atmosphere in the internodal lacunae of submerged sections, namely 3% O₂, 6% CO₂, 91% N₂ (by vol.) and 1 l l^-1 C₂H₄. Our results indicate that growth responses obtained with isolated rice stem sections are similar to those of intact deep-water rice plants.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - AVG aminoethoxyvinylglycine     

Formative and proliferative cell divisions,cell differentiation,and developmental changes in the meristem of Azolla roots

The root of the water fern Azolla is a compact higher-plant organ, advantageous for studies of cell division, cell differentiation, and morphogenesis. The cell complement of A. filiculoides Lam. and A. pinnata R.Br. roots is described, and the lineages of the cell types, all derived ultimately from a tetrahedral apical cell, are characterised in terms of sites and planes of cell division within the formative zone, where the initial cells of the cell files are generated. Subsequent proliferation of the initial cells is highly specific, each cell type having its own programme of divisions prior to terminal differentiation. Both formative and proliferative divisions (but especially the former) occur in regular sequences. Two enantiomorphic forms of root develop, with the dispositions of certain types of cell correlating with the direction, dextrorse or sinistrorse, of the cell-division sequence in the apical cells. Root growth is determinate, the apical cell dividing about 55 times, and its cell-cycle duration decreasing from an initial 10 h to about 4 h during the major phase of root development. Sites of proliferation progress acropetally during aging, but do not penetrate into the zone of formative divisions. The detailed portrait of root development that was obtained is discussed with respect to genetic and epigenetic influences; quantal and non-quantal cell cycles; variation in cell-cycle durations; relationships between cell expansion and cell division: the role of the apical cell; and the limitation of the total number of mitotic cycles during root formation.     

Hydroxyurea-induced mitotic synchronization in Allium sativum root meristems

The synchronized divisions following a treatment with hydroxyurea (HU) — an inhibitor of DNA synthesis — were studied in root meristems of Allium sativum using two methods: autoradiography of median sections and morphological labeling with a cytokinesis inhibitor. It is shown that the second wave of mitoses is heterogeneous: it is composed mostly of cells which have been synchronized in the S phase by the HU treatment, of cells coming from the quiescent center stimulated to enter DNA synthesis and of cells which were not blocked by the 23 h HU treatment (slow cycling cells). It is also shown that the cell cycle following the first synchronized division is considerably shortened by the synchronization procedure.Abbreviations QC quiescent center - HU hydroxyurea - MHQD methyl-3 hydroxy-6 quinazoline dione 2–4     

Mutations associated with floral organ number in rice

How floral organ number is specified is an interesting subject and has been intensively studied in Arabidopsis thaliana. In rice (Oryza sativa L.), mutations associated with floral organ number have been identified. In three mutants of rice, floral organ number 1 (fon1) and the two alleles, floral organ number 2-1 (fon2-1) and floral organ number 2-2 (fon2-2), the floral organs were increased in number centripetally. Lodicules, homologous to petals, were rarely affected, and stamens were frequently increased from six to seven or eight. Of all the floral organs the number of pistils was the most frequently increased. Among the mutants, fon1 showed a different spectrum of organ number from fon2 -1 and fon2 -2. Lodicules were the most frequently affected in fon1, but pistils of more than half of fon1 flowers were unaffected; in contrast, the pistils of most flowers were increased in fon2 -1 and fon2-2. Homeotic conversion of organ identity was also detected at a low frequency in ectopically formed lodicules and stamens. Lodicules and stamens were partially converted into anthers and stigmas, respectively. Concomitant with the increased number of floral organs, each mutant had an enlarged apical meristem. Although meristem size was comparable among the three mutants and wild type in the early phase of flower development, a significant difference became apparent after the lemma primordium had differentiated. In these mutants, the size of the shoot apical meristem in the embryo and in the vegetative phase was not affected, and no phenotypic abnormalities were detected. These results do not coincide with those for Arabidopsis in which clavatal affects the sizes of both shoot and floral meristems, leading to abnormal phyllotaxis, inflorescence fasciation and increased floral organs. Accordingly, it is considered that FON1 and FON2 function exclusively in the regulation of the floral meristem, not of the vegetative meristem.Abbreviation DIC differential interference contrast This work was supported in part by Grant-in-Aid for Scientific Research on Priority Areas from the Ministry of Education, Science and Culture of Japan.     

Effects of gibberellin on the cellular dynamics of dwarf pea internode development

This study analysed the dynamics of cell production and extension, and how these were affected by applied gibberellic acid (GA₃), during internode development in dwarf peas (Pisum sativum L. cv. Meteor). Image analysis was used to obtain cell number and length data for entire cell columns along the epidermis, the two outermost cortical layers, and the pith, from internode 7, over a time period covering the whole of the internode's growth phase. For a few days following the inception of an internode at the shoot apex, little further growth occurred, and there was no significant effect of GA₃ on cell division or cell extension. The subsequent growth of the internode was stimulated more than fourfold by GA₃ as a result of the production of more than twice the number of cells, which were twice as long. At least 96.5% of the cells of the mature internode were actually formed within the internode itself during this period of growth, demonstrating that the internode cells themselves represent the morphogenetic site of response to GA₃. Mitoses and cell extension occurred along the full length of the internode throughout its development. The daily changes in cell numbers were modelled by the Richards function, and manipulations of the fitted functions to reveal time trends of absolute and specific cell production rates were performed for each stem tissue. The increase in cell numbers in the +GA₃ plants was brought about by an increase in the rate of cell production, over a shorter time interval; specific cell production rates declined continuously from initial rapid rates in the +GA₃ epidermis and pith, but declined more slowly in the cortex. The control (−GA₃) epidermis and cortex cells exhibited a constant specific cell production rate (i.e. purely exponential) for several days. Cell extension rates were calculated so as to compensate for the size-reduction effects of concurrent cell division. These calculations confirmed that `real' cell extension rates were higher in the +GA₃ internodes. Models of the cellular controls of internode growth, based on the estimated dynamics of cell division and extension, are discussed. Received: 1 July 1997 / Accepted: 30 July 1997     

Immunochemical comparison of membrane-associated and secreted arabinogalactan-proteins in rice and carrot

Arabinogalactan-proteins (AGPs) occurring in suspension-cultured rice (Oryza saliva L.) cells, their conditioned medium and at the rice root apex were investigated using monoclonal antibodies and the AGP-binding -glucosyl Yariv reagent ( GlcY). A monoclonal antibody, LM2, was generated that recognized an acidic carbohydrate epitope common to two soluble AGPs occurring in the conditioned medium of proliferating rice cells, membrane-associated AGPs (rmAGP) in the cultured cells and two AGPs at the rice root apex. In addition, LM2 recognized AGPs secreted by suspensioncultured carrot (Daucus carota L.) cells. The two AGPs of the rice culture medium, srAGP1 and srAGP2, were discriminated by their mobilities during sodium dodecyl sulfate-polyacrylamide gel electrophoresis, reaction with GlcY, the presence of arabinogalactan epitopes and anion-exchange chromatography. The association of rmAGP with the plasma membrane was investigated by Triton-X-114/aqueous partitioning of both microsomal and plasma-membrane preparations and rmAGP was found to partition into the detergent phase, indicating that AGPs are hydrophobic plasma-membrane proteins in rice. This was in contrast to plasma-membrane AGPs of suspension-cultured carrot cells that partitioned into the aqueous phase. At the rice root apex most of the AGP was associated with the microsomal fraction and also partitioned into the detergent phase, although a distinct highmolecular-mass AGP entered the aqueous phase.Abbreviations AGP arabinogalactan-protein - GlcY -glucosyl Yariv reagent - ELISA enzyme-linked immunosorbent assay We gratefully acknowledge support from the Leverhulme Trust, the UK Biotechnology and Biological Sciences Research Council and the Royal Society.