Investigations concerning the cell cycle of Acer pseudoplatanus L. (sycamore) in suspension culture have generated the following general conclusions: 1) The sycamore suspensions used in the study exhibit extensive aneuploid development away from the diploid chromosome number of 2n = 4x = 52. 2) The evolution of new aneuploid chromosome complements may well still be in progress. 3) Stationary phase DNA distributions can be used to estimate chromosome numbers. 4) In common with animal and other plant cell types, the G1 phase is the most variable part of the DNA replication- partition cycle. 5) The S-G2-M sequence is of relatively constant duration in exponentially growing populations. 6) The S-G2-M sequence duration is lengthened in slowly dividing chemostat cultures, under glucose or nitrate limitation. 7) If cells are arrested in stationary phase by sucrose or phosphate starvation they arrest in the G1 and G2 phases in the approximate ratio of 4 : 1. If cells are arrested by nitrate starvation they accumulate almost exclusively in G1. 8) The G1 accumulation of nitrate starved cells is probably the mechanism by which synchrony is achieved when late stationary phase cells are inoculated into fresh medium. 9) Calculation of Scherbaum synchrony indices from cell count data, shows that the Acer system is well synchronised in terms of cell division. 10) The persistence of synchronous division and the absence of progressive synchrony decay, suggests that some form of intercellular entrainment occurs in the suspension cultures. 11) In synchronously dividing populations, DNA synthesis is also synchronous, and in terms of an S phase synchrony index, DNA replication becomes more highly synchronised as cell density rises and interphase duration is reduced. 12) Histones appear to be synthesised continuously during interphase in synchronised Acer suspensions but the rate of histone synthesis appears to be highest during S-phase.

Plant cell suspension cultures were used to study environmental factors that affected culture growth, greening and photosynthesis. Most experiments were conducted with cultures of Spinacia oleracea but some experiments involved Chenopodium giganteum or Atropa belladonna. Light (continuous white, 1.4 x 103 ergs.cm-2. sec-1 reflected irradiance) was demonstrated to be necessary for culture growth and greening in the absence of the organic growth factors of the medium. Cultures would grow in the dark in the presence of the organic growth factors (pages 67-86). Gas phase components were found to control growth and greening of lighted suspension cultures. (1) Reducing O2 from 21 to 5% AP promoted greening in conditions that prevented O2-limitation of growth. (2) Increasing CO2 from 5 to 10% inhibited culture growth and to a lesser extent greening. (3) Increasing C2H4 from 1 to 12 vpm caused a gradual parallel inhibition of culture growth and greening. Two sequential 336 h passages in 12 vpm C2H4 were toxic, resulting in cell death and chlorosis (pages 151-222). S, oleracea suspension cultures would grow and green in a salts plus sucrose (10 g 1-1) medium, based on that described by Murashige and Skoog (1964). 1-NAA (10-6M) applied in this medium promoted C2H4 production and chlorosis. Sucrose at any concentration between 0 and 20 g 1-1 inhibited greening, which was not overcome by glucose (20 g 1-1) substitution. Copper sulphate (10-6 M) was implicated as being involved in the protection mechanism against O2-toxicity (pages 87-150). Growth and greening of S, oleracea 1 to 5 1 cultures was inhibited by sparging, in contrast with rapidly stirred (vortex) gassed 1.5 1 cultures. The photosynthesis of S. oleracea cultures in certain cases equalled respiration rate (in terms of O2 exchange). Photosynthetic competence was related to chloroplast pigments content, the latter declining on subculture and reaching maximum values in early stationary phase of batch culture growth. Completely photo-autotrophic growth was not demonstrated.

The aim of this project was to study the accumulation of betalains in suspension cultures of <I>Beta vulgaris.</I> Betalains were found to accumulate in suspension cultures of <I>Beta vulgaris</I> in association with growth. The onset of pigment synthesis occurred prior to, or immediately after, the start of cell division. The production of betalains could be influenced by the conditions of culture differently from growth. Varying the concentration of inorganic nitrogen (i-N) or inorganic phosphorus (i-P); above or below that in B5 medium, did not increase either growth or pigment production. 3% sucrose was shown to be optimum for betalain accumulation while the fresh weight of culture was not affected. Lowering the concentration of 2,4-dichlorophenoxyacetic acid (2,4D) favoured betalain production. The highest pigment content per culture was achieved at 0.006 mg.1<SUP>-1</SUP> 2,4D at which concentration the culture growth was not significantly affected. Altering the concentration of kinetin in the medium did not increase the production of betalains. Administration of either tyrosine or dihydroxyphenylalanine (DOPA) did not significantly affect either growth or pigment production. A single cell plating techniques was used successfully to isolate a number of cell lines (violet, orange, yellow and white) with different qualitative and quantitative characteristics with regard to betacyanin and betaxanthin content. HPLC analysis of the betalains produced by the selected cell lines and the mother culture, revealed that betanin and miraxanthin V were the main betalains produced by the mother culture and the derived pigmented cell lines. These cell lines underwent changes in pigment production during a period of non-selective subculturing (15 months). The production of betalains in these cell lines was investigated in relation to cellular heterogeneity and the study showed that the cellular population of the cell lines contained a limited proportion of cells that accumulated the pigment.

The <I>Nicandra physaloides/Lycopersicon esculentum</I> heterograft consistently fails to produce functional vascular reconnections across the graft union (GU). Evidence supplied by earlier workers culminated in the hypothesis that a recognition system may exist between stock and scion. This thesis documents attempts to identify the stage [1] "off" signal for cell division in a model GU provided by actively-dividing suspension-cultured cells of <I>L.esculentum</I> x<I> peruvianum</I> and <I>L.esculentum</I> cv. Ailsa Craig (<I>L.esculentum</I> AC) by monitoring protein synthesis. The application of deproteinated cell walls of <I>N.physaloides, L.esculentum</I> AC, and <I>L.esculentum</I> x<I> peruvianum</I> consistently inhibited incorporation of [<SUP>14</SUP>C] leucine into protein, as did hemicelluloses from all sources at pH 4.5. pH measurements of apoplastic fluid from the GU of homografts and heterografts demonstrated a minimum value 4 d after grafting. In homografts GU pH increased thereafter, but in incompatible heterografts pH rose and then dropped. [<SUP>14</SUP>C]-labelled cell wall components were used to trace the fate of pectins and hemicelluloses applied to suspension-cultured cells; for cells of <I>L.esculentum</I> x <I>peruvianum</I> about 75% of the associated exogenous pectin was ionically bound to the cell surface, while 25% was internalised or bound by non-ionic means. At pH 6.0 about 73% of the associated hemicellulose bound to the cell surface while approximately 27% was internalised, but at pH 4.5 these proportions were almost 100% and 0% respectively. Experiments with cells of <I>L.esculentum</I> AC revealed that at pH 6.0 approximately 44% of the associated hemicellulose had bound to the cell surface, while about 55% had been internalised; when incubation occurred at pH 4.5 these percentages remained almost unchanged. The behaviour of the two types of suspension-cultured cells to identical stimuli is compared, and the relevance of the responses discussed in relation to graft compatibility.

The lens cell membranes and cell surface properties of two genotypes of chickens (HY-1 and HY-2) selected for high growth rate and associated with hyperplasia of the lens epithelium have been studied using a normal genotype as control. The results indicated that HY-1 and HY-2 cells are characterised by abnormalities in the structure and composition of their cell membranes. The membrane protein polypeptides differ markedly from each other both by charge and size. HY-1 and HY-2 membranes also have a high sialic acid content. Ultrastructural studies showed that both the intact epithelial cells and the membrane fractions of HY-1 and HY-2 cells have a marked deficiency of gap junctions. Immunological studies showed that although the membrane components of these cells share some antigenic properties with those of the normal, they are however not identical. Studies of lens epithelial cultures showed that these cells display numerous cell surface microvilli in contrast to their normal counterparts. It is suggested that this may be related to their increased lectin-binding and agglutinability, as revealed by studies with lectins, and their high metabolic rate as shown by their incorporation of radioactive precursors. Polysomes synthesising membrane components in these cells were studied by immunoprecipitation and the results revealed that these polysomes have a faster turnover than their normal counterparts. These cells bound more 45 calcium than normal cells but the activity of Na+-K+-ATPase in the lens epithelium of these cells was not significantly different from normal. It is suggested that the cell membrane modifications in these cells may be related to their abnormal cellular behaviour in cell culture and the hyperproliferation of the lens epithelium in vivo.

The aim of the work described in this thesis was to investigate the expression of mitochondrial, chloroplast and nuclear genes in successive sections along the length of the 7 day-old light-grown wheat leaf (Triticum aestivum). The wheat leaf represents a spatial separation of a temporal sequence of development, from basal meristematic to mature, photosynthetically competent cells. This sequence of cellular development is paralleled by a functional differentiation in which the energy requirement of the cell changes from an initial dependence upon oxidative phosphorylation to a combined and diumally modulated dependence on oxidative and photophosphoryla-tion. It was found that the abundance of specific mitochondrial genes (coxl, coxll, cob and atpA) per cell decreased between 3 and 10 fold within the basal lcm of the leaf and then remained at an approximately constant level to the distal tip. The abundance of specific mitochondrial RNA transcripts (coxl, coxll cob and AtpA) also decreased in relative abundance per cell beyond the basal lcm of the leaf, but in a manner which was not directly related to the decrease in mitochondrial gene copy number. Evidence was also found for the differential expression of nuclear gene(s) encoding the mitochondrial adenine nucleotide translocator. The relative abundance of specific mitochondrial proteins (the a-subunit of the Fi ATPase, subunit II of the cytochrome c oxidase complex, malate dehydrogenase and the adenine nucleotide translocator) per cell was found to remain approximately constant in the region to 4cm from the meristem. However the mitochondrial glycine decarboxylase protein was found to increase in relative abundance per cell in this region of the leaf. The abundance of specific chloroplast (rbcL and psbA) and nuclear (rbcS and cab) gene transcripts encoding chloroplast proteins were found to increase per cell along the length of the leaf from initially low levels in the meristematic region. These preliminary results illustrate the differential expression of nuclear and organelle genomes during cellular differentiation and indicate the complexity of the coordinated interactions required to satisfy the changing energy requirement of the plant cell.

Methods were developed for organogenesis and somatic embryogenesis of pistachio, <I>Pistacia vera </I>L. cv 'Antep', using tissues from seedlings, mature trees, immature fruits, zygotic embryos and juvenile leaf explants. A method was described for the establishment of embryonic cell suspension cultures. A procedure for the activation of meristem tips taken from 50-year old mature trees was also established. The factors controlling the initiation, maturation, germination, embling development, and acclimatisation of emblings derived from immature fruit explants were investigated. The induction of EMS was dependent on the type and concentration of PGRs and on the type of explants employed. The cytokinin BAP was found to be essential for the induction of EMS from immature fruits cultured on a liquid MS medium. The effects of various carbohydrate sources on the embryogenic capacity of EMS were examined. The best growth in terms of fresh and dry weight production was obtained on sucrose or glucose within the concentration range 4-10% w/v. Somatic embryos were found to mature more rapidly in liquid medium. An original method of logistic analysis was developed for interpretation of the effects of multiple treatments and their interactions on the probabilities of embryo germination and embling development. The abscisic acid and benzylaminopurine concentrations, the durations of the embryo maturation treatments and of the culture period for germination and embling development showed the most significant effects on embryo germination and embling development, which were higher when mature somatic embryos were transferred onto germination medium in clusters than as individuals.

The initiation and maintenance of cell division in Jerusalem Artichoke tubor tissue has been investigated in 2 different ways: 1. A kinetic analysis of ribosomal RNA synthesis and maturation was undertaken to determine the points where accumulation of this RNA is regulated. The evidence presented indicates that the massive increase in the level of RNA in response to auxin is in part due to an enhancement in transcription of the precursor rRNA and in part due to pout-transcriptional control involving an increase in the rates and degree of processing of the precursor to the mature rRNA. 2. The acidic proteins of the nucleus and the cytoplasm were investigated during the early stages of culture, prior to the onset of coil division in the auxin treated tissue. Both the synthesis of these proteins and their modification by phosphorylation were monitored by the highly discriminating 2-dimensional gel electrophoresis technique. Significant changes in the acidic proteins of the nucleus were found early in culture, with approximately 6 additional proteins being synthesized and accumulated in this organelle. As 0-1 phase progressed, the number was increased to 20 and then to 40 at the start of DNA replication. Alterations in the phosphorylation pattern occurred only at S, with the specific modification of 6 proteins. No changes in the cytoplasmic proteins were detected.

No description available.

No description available.