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The present work deals with a systematic investigation of 45 taxa belonging to 23 genera of the tribes Arabideae, Euclidieae, Hesperideae, Lunarieae, Matthioleae and Sisymbrieae of the family Brassicaceae from Egypt. This work is largely based on herbarium material received on loan from B, BM, BR, E, HUJ, K, KSU, L, LY, NMGM, OXF & FHO and W, and collections kept in the following herbaria; B, CAI, CAIM, WAG and Sohag University Herbarium (SHG, proposed abbreviation). In addition, fresh material of most of the taxa was studied and field observations were made from several localities in Egypt.Chapter 1 describes the family as a whole, presenting a historical review, its phytogeography, uses and economic importance, an outline of the thesis, aims and general objectives.Chapter 2 deals with the materials and methods used in this chapter. A full discussion for all morphological characters is provided; systematic treatment, based on macro-characters such as habit, leaf, floral, fruit, embryo and seed and micro-characters of trichomes. An artificial key applies the most reliable characters for the distinction of the genera.In the systematic treatment 45 taxa are recognized, presenting up-to-date nomenclature, literature citations, type specimens and their locations, synonyms, specimens examined, and distribution maps.Some taxonomic and nomenclature problems in Rorippa, Neslia, Eremobium, Malcolmia, Matthiola, Arabidopsis and Neotorularia are discussed and changes proposed.Chapter 3 presents macro-morphological characters of the seeds such as seed shape, colour, size, radicle, and micro- morphological characters by (SEM examination) of the epidermal cell shape, anticlinal boundaries, the outer periclinal cell wall and relief of outer cell walls are presented. Three types of basic anticlinal cell wall boundaries are recognized and, six different shapes of the outer periclinal cell wall are described. A key for the identification of the investigated taxa based on seed characters is provided.Chapter 4 deals with morphological characters of the pollen grains using SEM. In general the pollen morphology varies within a narrow range. The pollen grains are tricolpate. The shape varies from prolate spheroidal, subprolate to prolate. Three pollen types can be distinguished based on the size of lumina. The exine ornamentation of pollen grains varies between genera within tribes and between species within the same genus. The exine ornamentation of pollen was found useful to distinguish between closely related genera.Chapter 5 deals with the systematics of the tribes of the Brassicaceae, by means of numerical analysis based on sixty-two morphological characters, including vegetative parts, pollen grains and seeds. Morphological characters have been analyzed using two types of analyses: firstly, we performed a cluster analysis using average taxonomic distance and UPGMA clustering (procedures SIMINT, SAHN, and TREE). Secondly, we performed a principal coordinates analysis (PCO), using the product-moment correlation as a coefficient. The procedures STAND and SIMINT were used to calculate the distance matrix, the procedures EIGEN, PROJ, and MXPLOT to perform the PCO. Four main groups are recognised: Lunarieae, Euclidieae, Matthioleae and a mixed group. Representatives of these groups cluster together based on characters with high factor loading in the PCO analysis. The tribe Euclidieae is the most homogeneous group, and the tribe Arabideae is the most heterogeneous and probably of polyphyletic origin.Chapter 6 deals with phylogenetic relationships of 83 species belonging to 39 genera of tribes Brassiceae, Alysseae, Arabideae, Euclidieae, Hesperideae, Lepidieae, Lunarieae, Matthioleae and Sisymbrieae. In total 44 species belonging to 28 genera from Egypt were investigated using nucleotide sequences variation of the chloroplast matK gene to assess relationships among genera within tribe and tribes within the family. The topology of Jackknife, parsimony analysis, and Maximum Likelihood trees are largely in agreement. The analysis indicates that tribes Arabideae, Sisymbrieae, Hesperideae, and Lepidieae are polyphyletic.

De meeste pruimen die in Nederland geteeld worden, zijn nog altijd de rassen de Opal en Reine Victoria. Beide rassen zijn zelfbestuivers en hebben dus geen vreemd stuifmeel nodig voor een succesvolle zetting. Die zetting is bij deze rassen doorgaans zo sterk dat dunning een aanzienlijk deel van de teelt uitmaakt.

Voedseltekort oorzaak afname aantal wilde bijen. Vooral grote bijen zijn de dupe van gebreken in menu.

As a result of drying, the cytoplasm of desiccation-tolerant organisms, such as seed and pollen, enters into a highly viscous, solid-like, semi-equilibrium state: the glassy state. The work in this dissertation is focussed on the function and characteristics of intracellular glasses in these organisms.It was established that intracellular glasses are formed in both desiccation-tolerant and -intolerant pollen (chapter 1). However, desiccation-intolerant pollen loses its viability during drying before intracellular glasses are formed. This indicates that desiccation tolerance is not related with the formation of glasses during drying. Storage of cattail ( Typha latifolia ) pollen under different water contents and temperatures revealed the existence of an optimum water content for survival at a constant relative humidity (11-15%) (chapter 2). The water content corresponding to this relative humidity shifted to higher values with lower storage temperatures, and was found to be associated with the Brunauer, Emmet, and Teller monolayer value. Drying of the pollen below these water contents had detrimental effects on longevity. The water content-temperature combinations of optimal storage were found to be below the glass transition curve, indicating that optimum storage conditions are achieved when intracellular glasses are present. There was a change in ageing kinetics of cattail pollen associated with the melting of the intracellular glass. Above the glass transition temperature (T g ) the activation energy of the ageing rates increased two to three times. This suggests that the presence of glasses in the dry state improves storage stability by decreasing viscosity and, thus, ageing rate. It was concluded that T g curves might be useful for predictions of storage longevity above optimum water contents. However, they cannot be used solely to predict the precise conditions of optimum storage. Subsequently, we sought for a more direct measurement to assess the viscosity of the cytoplasm of tissues.For this purpose, we used electron paramagnetic resonance (EPR) spectroscopy to study the molecular mobility of the hydrophilic nitroxide spin probe 3-carboxy-proxyl (CP) that was incorporated into embryonic axes of pea seeds and cattail pollen. Using the distance between the outer extrema of the EPR spectrum (2 A zz ) as a measure of molecular mobility, a sharp increase in mobility was observed at a definite temperature during heating (chapter 3). This temperature increased with decreasing water content of the samples, and was found to be associated with the melting of the glassy state as measured by differential scanning calorimetry (DSC). Molecular mobility was found to be inversely correlated with storage stability: the higher the molecular mobility, the shorter the longevity: with decreasing water content, the molecular mobility reached a minimum, in concert with ageing rates. At very low water contents, both molecular mobility and ageing rates increased again. Minimum mobility and maximum storage stability occurred at similar water contents, suggesting that storage stability might be partially controlled by molecular mobility. To understand the nature of the changes in 2 A zz in spectra of CP in the tissues, echo detected (ED) EPR spectroscopy was employed (chapter 4). The shape of the ED EPR spectrum revealed the presence of librational motion of the spin probe, a motion typically present in glassy materials. The change in 2 A zz appeared to be the result of librational motion of the spin probe.With the use of saturation transfer (ST) EPR spectroscopy, a more quantitative measure of molecular mobility was acquired: the rotational correlation time (τ R ), which corresponds to the time it takes for the spin probe to rotate a radian around its axis (chapter 5). At room temperature,τ R of CP in pea embryonic axes increased during drying from 10 -11 s in de hydrated state to 10 -4 s in the dry state. At T g ,τ R was constant at10 -4 s for all water contents studied. The temperature dependence ofτ R at all water contents studied followed Arrhenius behaviour with a break at T g . The temperature effect onτ R above T g was much smaller in pea axes as found previously for sugar and polymer glasses. Thus, the melting of the intracellular glass by raising the temperature caused only a moderate increase in molecular mobility in the cytoplasm as compared to a huge increase in amorphous sugars.The application of saturation transfer EPR spectroscopy to biological tissues enabled a quantitative comparison between storage stability and molecular mobility in different tissues (section III). The temperature and moisture dependence of ageing rates of seeds and pollen was found to correlate with the rotational motion of CP in the cytoplasm (chapter 6-8). An increase in the temperature resulted in a faster rotational motion in the cytoplasm of cattail pollen, analogous to faster ageing rates (chapter 6). Decreasing the water content of the pollen resulted in a decrease in rotational motion until a minimum was reached, after which rotational motion slightly increased again. The water content at which this minimal rotational motion was observed increased with decreasing temperature, comparable to the pattern of ageing rate. A significant linear relationship was found between ageing rates and rotational motion in the cytoplasm of the pollen.We also investigated the relationship between the longevity of lettuce seeds and the molecular mobility in the cytoplasm of their radicles (chapter 7). Longevity of lettuce seeds was predicted using the viability equation of Ellis and Roberts. Increasing the temperature resulted in faster rotational motion and shorter longevity. There was a linear relationship between the logarithms of rotational motion and estimated longevity for temperatures above 5°C, which is the same temperature range for which experimental data were used to obtain the viability constants of the viability equation. Below 5°C, there was a deviation from linearity, which might stem from inaccurate predictions by the viability equation at low temperatures.Chapter 8 further demonstrates that there is a linear relationship between the logarithms of rotational motion in the cytoplasm of seed and pollen of several plant species and their ageing rates or longevities. This linearity suggests that cytoplasmic mobility might be an important controlling factor of ageing rates. The linear relationship between the two parameters could be used to predict lifespan of germplasm at low temperatures (at which experimental determination of longevity is practically impossible) by simply measuring theτ R values at these low temperatures (chapter 7 and 8). Based on the predictions using the linear regression between ageing rate and rotational motion of CP in pea embryonic axes, an optimum water content of storage was found. This optimum water content shifted to higher values with lowering the storage temperature, as was found previously for cattail pollen based on experimental data (chapter 2). It was predicted that the longevity of seeds at high (0.12 to 0.16 g/g) water content is much higher than previously suggested on the basis of the viability equation. The predictions show that drying germplasm too far leads to decreased longevity compared to storage of germplasm at higher water contents, suggesting that current storage protocols might have to be re-examined.Desiccation-tolerant organisms contain large amounts of soluble sugars. This, and the fact that sugars are excellent glass-formers has led to the suggestion that sugars play an important role in intracellular glass formation. The presence and amounts of oligosaccharides have been found to correlate with longevity. Furthermore, oligosaccharide glasses are known to increase the T g and viscosity of model sucrose glasses. This suggests that oligosaccharides might enhance the stability of intracellular glasses (chapter 9 and 10). Osmo-priming, i.e. pre-imbibition of seeds in an osmotic solution, can result in a decrease in oligosaccharide content and longevity. Priming pea seeds decreased the total oligosaccharide content in the embryonic axes (chapter 9). Despite the change in oligosaccharide:sucrose ratio, no differences in T g values were detected in the dry axes before and after priming as determined by DSC. Also no difference was found between the rotational mobility of CP in dry untreated axes and that of dry primed axes. Chapter 10 demonstrates that osmo-priming of impatiens and bell pepper seeds resulted in considerable decreases in longevity and oligosaccharide contents, while sucrose contents increased. Again, no differences in the T g curves were found between control and primed impatiens seeds. Similarly, there was no difference in rotational motion of CP in the cytoplasm between control and primed impatiens seeds and between control and primed bell pepper embryonic axes. It was concluded that oligosaccharides in seeds do not appear to affect the stability of the intracellular glassy state, and that the reduced longevity after priming is not the result of increased molecular mobility in the cytoplasm.To understand the nature and composition of biological glasses we investigated the molecular mobility around T g in sugars, poly-L-lysine and dry desiccation-tolerant biological systems, using ST-EPR, 1 H-NMR and FTIR spectroscopy. Two distinct changes in the temperature dependence of molecular mobility were detected in sugars and poly-L-lysine. With heating, the first change was associated with the melting of the glassy state (T g ). The second change, at which the molecular mobility abruptly increased over several orders of magnitude, was found to correspond with a critical temperature (T c ) where the dynamics of the system changed from solid-like to liquid-like. The temperature interval between T g and T c increased with increasing molecular weight of the sugars. The interval between T g and T c in biological tissues was over 50°C, implying that the stability remained high even at temperatures far above T g . A comparably high T c -T g interval was found for the molecular mobility of poly-L-lysine, suggesting that proteins rather than sugars play an important role in the intracellular glass formation. The exceptionally high T c of intracellular glasses is expected to provide excellent long-term stability to dry organisms, maintaining a slow molecular motion in the cytoplasm even at temperatures far above T g .

As a result of drying, the cytoplasm of desiccation-tolerant organisms, such as seed and pollen, enters into a highly viscous, solid-like, semi-equilibrium state: the glassy state. The work in this dissertation is focussed on the function and characteristics of intracellular glasses in these organisms.It was established that intracellular glasses are formed in both desiccation-tolerant and -intolerant pollen (chapter 1). However, desiccation-intolerant pollen loses its viability during drying before intracellular glasses are formed. This indicates that desiccation tolerance is not related with the formation of glasses during drying. Storage of cattail ( Typha latifolia ) pollen under different water contents and temperatures revealed the existence of an optimum water content for survival at a constant relative humidity (11-15%) (chapter 2). The water content corresponding to this relative humidity shifted to higher values with lower storage temperatures, and was found to be associated with the Brunauer, Emmet, and Teller monolayer value. Drying of the pollen below these water contents had detrimental effects on longevity. The water content-temperature combinations of optimal storage were found to be below the glass transition curve, indicating that optimum storage conditions are achieved when intracellular glasses are present. There was a change in ageing kinetics of cattail pollen associated with the melting of the intracellular glass. Above the glass transition temperature (T g ) the activation energy of the ageing rates increased two to three times. This suggests that the presence of glasses in the dry state improves storage stability by decreasing viscosity and, thus, ageing rate. It was concluded that T g curves might be useful for predictions of storage longevity above optimum water contents. However, they cannot be used solely to predict the precise conditions of optimum storage. Subsequently, we sought for a more direct measurement to assess the viscosity of the cytoplasm of tissues.For this purpose, we used electron paramagnetic resonance (EPR) spectroscopy to study the molecular mobility of the hydrophilic nitroxide spin probe 3-carboxy-proxyl (CP) that was incorporated into embryonic axes of pea seeds and cattail pollen. Using the distance between the outer extrema of the EPR spectrum (2 A zz ) as a measure of molecular mobility, a sharp increase in mobility was observed at a definite temperature during heating (chapter 3). This temperature increased with decreasing water content of the samples, and was found to be associated with the melting of the glassy state as measured by differential scanning calorimetry (DSC). Molecular mobility was found to be inversely correlated with storage stability: the higher the molecular mobility, the shorter the longevity: with decreasing water content, the molecular mobility reached a minimum, in concert with ageing rates. At very low water contents, both molecular mobility and ageing rates increased again. Minimum mobility and maximum storage stability occurred at similar water contents, suggesting that storage stability might be partially controlled by molecular mobility. To understand the nature of the changes in 2 A zz in spectra of CP in the tissues, echo detected (ED) EPR spectroscopy was employed (chapter 4). The shape of the ED EPR spectrum revealed the presence of librational motion of the spin probe, a motion typically present in glassy materials. The change in 2 A zz appeared to be the result of librational motion of the spin probe.With the use of saturation transfer (ST) EPR spectroscopy, a more quantitative measure of molecular mobility was acquired: the rotational correlation time (τ R ), which corresponds to the time it takes for the spin probe to rotate a radian around its axis (chapter 5). At room temperature,τ R of CP in pea embryonic axes increased during drying from 10 -11 s in de hydrated state to 10 -4 s in the dry state. At T g ,τ R was constant at10 -4 s for all water contents studied. The temperature dependence ofτ R at all water contents studied followed Arrhenius behaviour with a break at T g . The temperature effect onτ R above T g was much smaller in pea axes as found previously for sugar and polymer glasses. Thus, the melting of the intracellular glass by raising the temperature caused only a moderate increase in molecular mobility in the cytoplasm as compared to a huge increase in amorphous sugars.The application of saturation transfer EPR spectroscopy to biological tissues enabled a quantitative comparison between storage stability and molecular mobility in different tissues (section III). The temperature and moisture dependence of ageing rates of seeds and pollen was found to correlate with the rotational motion of CP in the cytoplasm (chapter 6-8). An increase in the temperature resulted in a faster rotational motion in the cytoplasm of cattail pollen, analogous to faster ageing rates (chapter 6). Decreasing the water content of the pollen resulted in a decrease in rotational motion until a minimum was reached, after which rotational motion slightly increased again. The water content at which this minimal rotational motion was observed increased with decreasing temperature, comparable to the pattern of ageing rate. A significant linear relationship was found between ageing rates and rotational motion in the cytoplasm of the pollen.We also investigated the relationship between the longevity of lettuce seeds and the molecular mobility in the cytoplasm of their radicles (chapter 7). Longevity of lettuce seeds was predicted using the viability equation of Ellis and Roberts. Increasing the temperature resulted in faster rotational motion and shorter longevity. There was a linear relationship between the logarithms of rotational motion and estimated longevity for temperatures above 5°C, which is the same temperature range for which experimental data were used to obtain the viability constants of the viability equation. Below 5°C, there was a deviation from linearity, which might stem from inaccurate predictions by the viability equation at low temperatures.Chapter 8 further demonstrates that there is a linear relationship between the logarithms of rotational motion in the cytoplasm of seed and pollen of several plant species and their ageing rates or longevities. This linearity suggests that cytoplasmic mobility might be an important controlling factor of ageing rates. The linear relationship between the two parameters could be used to predict lifespan of germplasm at low temperatures (at which experimental determination of longevity is practically impossible) by simply measuring theτ R values at these low temperatures (chapter 7 and 8). Based on the predictions using the linear regression between ageing rate and rotational motion of CP in pea embryonic axes, an optimum water content of storage was found. This optimum water content shifted to higher values with lowering the storage temperature, as was found previously for cattail pollen based on experimental data (chapter 2). It was predicted that the longevity of seeds at high (0.12 to 0.16 g/g) water content is much higher than previously suggested on the basis of the viability equation. The predictions show that drying germplasm too far leads to decreased longevity compared to storage of germplasm at higher water contents, suggesting that current storage protocols might have to be re-examined.Desiccation-tolerant organisms contain large amounts of soluble sugars. This, and the fact that sugars are excellent glass-formers has led to the suggestion that sugars play an important role in intracellular glass formation. The presence and amounts of oligosaccharides have been found to correlate with longevity. Furthermore, oligosaccharide glasses are known to increase the T g and viscosity of model sucrose glasses. This suggests that oligosaccharides might enhance the stability of intracellular glasses (chapter 9 and 10). Osmo-priming, i.e. pre-imbibition of seeds in an osmotic solution, can result in a decrease in oligosaccharide content and longevity. Priming pea seeds decreased the total oligosaccharide content in the embryonic axes (chapter 9). Despite the change in oligosaccharide:sucrose ratio, no differences in T g values were detected in the dry axes before and after priming as determined by DSC. Also no difference was found between the rotational mobility of CP in dry untreated axes and that of dry primed axes. Chapter 10 demonstrates that osmo-priming of impatiens and bell pepper seeds resulted in considerable decreases in longevity and oligosaccharide contents, while sucrose contents increased. Again, no differences in the T g curves were found between control and primed impatiens seeds. Similarly, there was no difference in rotational motion of CP in the cytoplasm between control and primed impatiens seeds and between control and primed bell pepper embryonic axes. It was concluded that oligosaccharides in seeds do not appear to affect the stability of the intracellular glassy state, and that the reduced longevity after priming is not the result of increased molecular mobility in the cytoplasm.To understand the nature and composition of biological glasses we investigated the molecular mobility around T g in sugars, poly-L-lysine and dry desiccation-tolerant biological systems, using ST-EPR, 1 H-NMR and FTIR spectroscopy. Two distinct changes in the temperature dependence of molecular mobility were detected in sugars and poly-L-lysine. With heating, the first change was associated with the melting of the glassy state (T g ). The second change, at which the molecular mobility abruptly increased over several orders of magnitude, was found to correspond with a critical temperature (T c ) where the dynamics of the system changed from solid-like to liquid-like. The temperature interval between T g and T c increased with increasing molecular weight of the sugars. The interval between T g and T c in biological tissues was over 50°C, implying that the stability remained high even at temperatures far above T g . A comparably high T c -T g interval was found for the molecular mobility of poly-L-lysine, suggesting that proteins rather than sugars play an important role in the intracellular glass formation. The exceptionally high T c of intracellular glasses is expected to provide excellent long-term stability to dry organisms, maintaining a slow molecular motion in the cytoplasm even at temperatures far above T g .

De eik houdt de genetische variatie op peil en voorkomt inteelt doordat de moedereik selecteert op stuifmeel van ver verwijderd staande eiken. Dit blijkt uit DNA-onderzoek van de eikels van groepjes eiken op locaties in het Leuvenumse bos en in Amsterdam

To study transcription and maturation of RNA, pollen development and androgenesis of Brassica napus was chosen as a test system as explained in Chapter 1. Culturing microspores and young bicellular pollen at 32 °C allows to change the developmental pathway from pollen development towards embryogenesis. Culturing at 18 °C results in normal pollen development.The ultrastructure of the nucleus was analyzed during pollen development applying various staining techniques which visualised the degree of chromatin condensation, the appearance of the interchromatin region and the nucleolar structure. The interchromatin region contains proteins which are necessary for the transcription and maturation of RNA.The results give a rough indication about the transcriptional activity at various pollen stages. Although difficult to interpret, they indicate an increase in transcription activity in nuclei from the young microspore stage up to the vegetative cell in almost mature pollen. The generative nucleus in early pollen initially showed all signs of a transcriptionally active nucleus, but it lost its transcriptionally active morphology prior to pollen mitosis. After pollen mitosis the chromatin of the sperm nuclei remained condensed, indicating that the sperm nuclei are transcriptionally silenced.To get more insight into the cell cycle stages of the various nuclei of bicellular and tricellular pollen and under embryogenic and non-embryogenic culture conditions, the DNA synthesis was analysed using bromodeoxyuridine. It was found that microspores are able to change their developmental pathway towards embryogenesis from G1 upto G2. In bicellular pollen the vegetative nucleus is arrested in the G1 and re-enters the cell cycle after embryo induction (Binarova et al., Theor. Appl. Genet. 87 : 9 - 16).To visualise RNA synthesis in plant nuclei in detail, procedures were developed to label the sites of transcription using the uridine-analogue 5'-bromouridine 5'-triphosphate (BrUTP). It is assessed that this method works well on isolated leaf cell nuclei of Brassica napus . It enabled to visualize transcription sites throughout the plant nucleus like was shown before in nuclei of vertebrate cells (Straatman et al., Protoplasma 192 : 145 - 149).The method was further developed, using leaf and pollen protoplasts, and germinating pollen with pollen tubes. In pollen protoplasts and pollen tubes, labelling of RNA transcription was only found in the vegetative nucleus; the sperm nuclei were silenced, which is in agreement with the ultrastructural analysis. It was further found that transcription in the vegetative nucleus of mature pollen protoplasts could be very intense whereas the transcription of this nucleus was relatively low in pollen tubes. However, attempts to label transcription sites during pollen development and after embryo-induction did not result in any signal.Leaf and pollen protoplasts were also used to analyse the maturation of pre-mRNA and to understand the relation between the transcription sites and the location of small nuclear ribonucleoprotein particles (snRNPs) and small nucleolar RNPs (snoRNPs). The snRNP distribution was also studied during pollen development and androgenesis. snRNPs were not detected in the generative nucleus of late bicellular pollen and in the sperm nuclei of tricellular pollen. This is in accordance with the ultrastructural analysis of these stages where the chromatin was more condensed in late bicellular pollen than in earlier stages indicating a low or silenced transcription. In sperm cells the chromatin was even more condensed and no incorporation of BrUTP was detected. However, also the vegetative nucleus of mature pollen did not label with antibodies against snRNPs, although the chromatin was very dispersed which points to a high transcriptional activity, as was found in the pollen protoplasts using BrUTP.A coiled body is one of the subcompartments in the nucleus containing snRNPs and fibrillarin. In general, the number of coiled bodies in nuclei of mammalian cells increases with an increase in transcriptional activity. Although there was an increase in the number of coiled bodies, the increase in size of these bodies was much more dramatic during pollen development. After embryo induction a slow decrease in size of the coiled bodies was found.All RNA leaving the nucleus and all nuclear proteins produced in the cytoplasm have to pass the nuclear pore complexes (NPCs). Therefore, the NPC density can give an indication about general nuclear activity. During pollen development the NPC density slowly increased and reached its highest level in the vegetative nucleus of mature pollen. This correlates well with the fact that just before the vegetative nucleus is silenced, the transcription is high in this nucleus. The NPC density of the sperm nuclei was low which confirms the state of low activity as has been described. After or during embryo induction a decrease in NPC density was found. However, this occurred under both embryogenic and non-embryogenic conditions, indicating that it is not evidence for embryo induction.

To study transcription and maturation of RNA, pollen development and androgenesis of Brassica napus was chosen as a test system as explained in Chapter 1. Culturing microspores and young bicellular pollen at 32 °C allows to change the developmental pathway from pollen development towards embryogenesis. Culturing at 18 °C results in normal pollen development.The ultrastructure of the nucleus was analyzed during pollen development applying various staining techniques which visualised the degree of chromatin condensation, the appearance of the interchromatin region and the nucleolar structure. The interchromatin region contains proteins which are necessary for the transcription and maturation of RNA.The results give a rough indication about the transcriptional activity at various pollen stages. Although difficult to interpret, they indicate an increase in transcription activity in nuclei from the young microspore stage up to the vegetative cell in almost mature pollen. The generative nucleus in early pollen initially showed all signs of a transcriptionally active nucleus, but it lost its transcriptionally active morphology prior to pollen mitosis. After pollen mitosis the chromatin of the sperm nuclei remained condensed, indicating that the sperm nuclei are transcriptionally silenced.To get more insight into the cell cycle stages of the various nuclei of bicellular and tricellular pollen and under embryogenic and non-embryogenic culture conditions, the DNA synthesis was analysed using bromodeoxyuridine. It was found that microspores are able to change their developmental pathway towards embryogenesis from G1 upto G2. In bicellular pollen the vegetative nucleus is arrested in the G1 and re-enters the cell cycle after embryo induction (Binarova et al., Theor. Appl. Genet. 87 : 9 - 16).To visualise RNA synthesis in plant nuclei in detail, procedures were developed to label the sites of transcription using the uridine-analogue 5'-bromouridine 5'-triphosphate (BrUTP). It is assessed that this method works well on isolated leaf cell nuclei of Brassica napus . It enabled to visualize transcription sites throughout the plant nucleus like was shown before in nuclei of vertebrate cells (Straatman et al., Protoplasma 192 : 145 - 149).The method was further developed, using leaf and pollen protoplasts, and germinating pollen with pollen tubes. In pollen protoplasts and pollen tubes, labelling of RNA transcription was only found in the vegetative nucleus; the sperm nuclei were silenced, which is in agreement with the ultrastructural analysis. It was further found that transcription in the vegetative nucleus of mature pollen protoplasts could be very intense whereas the transcription of this nucleus was relatively low in pollen tubes. However, attempts to label transcription sites during pollen development and after embryo-induction did not result in any signal.Leaf and pollen protoplasts were also used to analyse the maturation of pre-mRNA and to understand the relation between the transcription sites and the location of small nuclear ribonucleoprotein particles (snRNPs) and small nucleolar RNPs (snoRNPs). The snRNP distribution was also studied during pollen development and androgenesis. snRNPs were not detected in the generative nucleus of late bicellular pollen and in the sperm nuclei of tricellular pollen. This is in accordance with the ultrastructural analysis of these stages where the chromatin was more condensed in late bicellular pollen than in earlier stages indicating a low or silenced transcription. In sperm cells the chromatin was even more condensed and no incorporation of BrUTP was detected. However, also the vegetative nucleus of mature pollen did not label with antibodies against snRNPs, although the chromatin was very dispersed which points to a high transcriptional activity, as was found in the pollen protoplasts using BrUTP.A coiled body is one of the subcompartments in the nucleus containing snRNPs and fibrillarin. In general, the number of coiled bodies in nuclei of mammalian cells increases with an increase in transcriptional activity. Although there was an increase in the number of coiled bodies, the increase in size of these bodies was much more dramatic during pollen development. After embryo induction a slow decrease in size of the coiled bodies was found.All RNA leaving the nucleus and all nuclear proteins produced in the cytoplasm have to pass the nuclear pore complexes (NPCs). Therefore, the NPC density can give an indication about general nuclear activity. During pollen development the NPC density slowly increased and reached its highest level in the vegetative nucleus of mature pollen. This correlates well with the fact that just before the vegetative nucleus is silenced, the transcription is high in this nucleus. The NPC density of the sperm nuclei was low which confirms the state of low activity as has been described. After or during embryo induction a decrease in NPC density was found. However, this occurred under both embryogenic and non-embryogenic conditions, indicating that it is not evidence for embryo induction.