Segregation of genetic markers among plants regenerated from cultured anthers of broccoli (Brassica oleracea var. ‘italica’)

Summary An experiment was conducted to determine critical factors in the recovery of embryos from cultured anthers of broccoli (Brassica oleracea var. italica) and to unambiguously distinguish whether embryos were of gametophytic origin. Among factors tested, genotype, genotype x anther developmental stage, and method of anther culture had a distinct impact on embryo recovery, whereas length of anther exposure to the culture medium did not. However, extreme heterogeneity of embryo emergence within and among replications precluded statistical contrasts. Among 762 plants derived from embryos of four independent cultivars, only one was determined to be of sporophytic origin by use of heterozygous codominant isozyme markers. Two of the cultivars tested were heterozygous at two or more loci. While segregation among loci was consistent with previously published linkage data, segregation of alleles was consistently non-random. In all of seven separate cases involving four cultivars, a significant over-representation of the fast-migrating class was observed. It appears, therefore, that populations of plants derived from microspores within cultured anthers of broccoli do not necessarily represent a random gametic array, and that care must be exercised in breeding and genetic applications.     

Antioxidant activity and ROS tolerance in triticale (×Triticosecale Wittm.) anthers affect the efficiency of microspore embryogenesis

To verify the hypothesis that cell redox status regulates the process of microspore embryogenesis (ME), reactive oxygen species (ROS) generation and the activity of enzymatic and non-enzymatic antioxidants were analyzed in eight doubled haploid lines of triticale with significant differences in embryogenic potential. The analyses were performed in anthers excised from freshly cut tillers (control) and from low temperature (LT) pre-treated tillers (3 weeks at 4 °C) in which ME has been initiated. Significant associations between ME effectiveness and the variables studied were found. In control cultures, high superoxide dismutase (SOD) activity appeared crucial for microspore viability. On the other hand, positive though non-linear correlation between ME effectiveness and H₂O₂ generation, and negative correlation with catalase (CAT) activity suggest that some threshold level of H₂O₂ is important for successful ME initiation. LT tillers pre-treatment significantly increased H₂O₂ accumulation, which had a negative effect on ME effectiveness. However, even high level of H₂O₂ did not endanger cell viability as long as the cells exhibited high activity of ROS-decomposing enzymes (SOD, CAT and POX). The ability to sustain antioxidative enzyme activity under cold stress in the dark was another important requirement for high effectiveness of ME, allowing for the generation of the signal initiating microspore reprogramming and simultaneously protecting the cells from the toxic effects of ROS production. The role of antioxidative enzymes cannot be replaced even by high activity of non-enzymatic antioxidants. In conclusion, genetically controlled but environmentally modified cell tolerance to oxidative stress seems to play an important role in triticale ME.     

We appreciate the invaluable contributions of the following reviewers during the editing of Vol. 8 Nos. 1–2.

Acknowledgments

We appreciate the invaluable contributions of the following reviewers during the editing of Vol. 8 Nos. 1–2.     

Population-level variation in the expression of heterostyly in three species of Rubiaceae: does reciprocal placement of anthers and stigmas characterize heterostyly?

Heterostyly (i.e., reciprocal placement of anthers and stigmas between two or three floral morphs) is hypothesized to enhance outcrossing and reduce selfing. However, few studies have documented reciprocity among individual plants; instead, mean anther and stigma heights for floral morphs are usually reported, masking interindividual variation. We measured eight floral dimensions for individuals in five populations of three heterostylous Rubiaceae. The three methods used to quantify reciprocity yielded different conclusions regarding the degree to which populations conformed to expectations for heterostylous plants. Only Psychotria poeppigiana had stigma and, to a lesser degree, anther heights in discrete classes. Variation among plants of Bouvardia ternifolia and Psychotria chiapensis yielded a continuum of anther and stigma heights across populations. Comparison of distances between stigma and anthers indicated that only flowers of B. ternifolia had, as expected, a constant value for this distance. Finally, regression relationships between anther and stigma heights and corolla length showed that only in one population each of B. ternifolia and P. poeppigiana, and in P. chiapensis, was distance between anthers and stigmas the same across the range of corolla sizes for both floral morphs. Variation among these species in expression of heterostyly was not clearly linked to phylogenetic relationship or pollinator syndromes. Two approach herkogamous (AH) species were studied for comparison. Flowers of Psychotria brachiata were consistently AH, but flowers of P. pittieri were highly variable. Determining fitness consequences of population-level variation in sexual systems requires studies linking floral morphology to pollinator behavior and pollen transfer.     

Ecology of the pteridophytes on the southern slopes of Mt. Kilimanjaro – I. Altitudinal distribution

140 taxa of 61 genera in 24 families of pteridophytes were recorded on the southern slopes of Mt. Kilimanjaro. These represent about one third of the entire pteridophyte flora of Tanzania. The families richest in species are the Aspleniaceae, the Adiantaceae, the Dryopteridaceae, the Thelypteridaceae and the Hymenophyllaceae. Due to its luxuriant montane rain forest, which receives a precipitation of up to over 3000 mm, Mt. Kilimanjaro is distinctly richer in pteridophyte species than other volcanoes in East Africa. However, compared with the mountains of the Eastern Arc, the number of pteridophytes on Mt. Kilimanjaro is smaller. This can be explained by the widely destroyed submontane (intermediate) forest rather than by the higher age of the Eastern Arc Mts.The altitudinal distribution of all ferns was investigated in 24 transects. On the southern slopes of Mt. Kilimanjaro they were found in an altitudinal range of 3640 m. Cyclosorus quadrangularis, Azolla nilotica, Azolla africana andMarsilea minuta are restricted to the foothills, while Polystichum wilsonii, Cystopteris nivalis and Asplenium adiantum-nigrum are species found in the highest altitudes.Based on unidimensionally constrained clustering and on the analysis of the lowermost and uppermost occurrence of species, floristic discontinuities within the transects were determined. From these data and from an evaluation of the distribution of ecological groups and life forms, 11 altitudinal zones could be distinguished: a colline zone (–900 m asl), a submontane zone (900–1600 m asl) with lower and upper subzones, a montane zone (1600-2800 m asl) divided into 4 subzones, a subalpine zone (2800–3900 m asl) with lower, middle and upper subzones, and finally a (lower) alpine zone above 3900 m. The highest species numbers were observed in the lower montane forest belt between 1600 and 2000 m altitude. The zonation of ferns found at Mt. Kilimanjaro corresponds well with the vegetational zonation described by other authors using bryophytes as indicators in different parts of the humid tropics.     

Chemical agents that inhibit pollen development: effects of the phenyl-cinnoline carboxylates SC-1058 and SC-1271 on the ultrastructure of developing wheat anthers (Triticum aestivum L. var Yecora rojò)

Summary Phenylcinnoline carboxylate compounds SC-1058 and SC-1271 cause complete male sterility in wheat when applied at suitable dosages at the pre-meiotic stage of anther development. Anthers from treated and untreated plants were compared using light and electron microscopy from the pre-meiotic stage through the formation of nearly mature pollen. Overall anther development is gradually slowed in treated plants and pollen development is generally arrested in the late prevacuolate or early vacuolate microspore stage, although the first pollen mitosis does sometimes occur. The sporopollenin-containing exine walls are thinner, and show abnormally developed foot and tectum layers with sparse connecting baculi. Microspore cytoplasm degenerates and the cells eventually collapse. At the early, prevacuolate, free microspore stage treated tapetal cells hypertrophy, expanding into the locule. They contain abnormally large vacuoles that appear to form from the fusion of secretory vesicles, and some vacuoles contain electrondense deposits. The sporopollenin-containing orbicular wall and Ubisch bodies are retarded in their development and are structurally deformed. Acetolysis of whole anthers and of thick sections shows that the sporopollen-in-containing structures of treated materials are greatly reduced in thickness and are less rigid than in the control. We conclude that application of these compounds causes interference with the secretory function of tapetal cells which supplies sporopollenin cell-wall polymers to the exine of the microspores and to the tapetal orbicular wall and associated Ubisch bodies. Interference with the tapetal secretion of other nutrients required for microspore development is strongly suggested.     

Interaction of α-lactalbumin with dimyristoylphosphatidylcholine vesicles. III. Influence of the temperature and of the lipid-to-protein molar ratio on the complex formation

We investigated the interaction between α-lactalbumin and sonicated dimyristoylphosphatidylcholine at pH 4 and different temperatures. (1) At 23°C and lipid-to-protein molar ratios below 170, the interaction results in a disruption of the original vesicles to form smaller complex particles. By the sedimentation velocity method we determined for this particle a molar mass of (1.05 ± 0.16) · 10⁶ g·mol^?1. The lipid-to-protein molar ratio within the complex particle is 70/1, as earlier estimated. It follows that there are approximately 1200 lipid and 17 α-lactalbumin molecules per particle. At molar ratios above 170, α-lactalbumin strongly associates with the vesicles. In this case the vesicle entity remains. The ability of α-lactalbumin to break up the vesicles at this temperature is determined by the number of protein molecules which are required in the complex particle. (2) By means of fluorescence polarization of the lipophilic probe 1,6-diphenyl-1,3,5-hexatriene and energy transfer of the tryptophan groups of the protein to 1,3-(1,1′-dipyrenyl)propane located in the hydrocarbon region of the vesicles, it is shown that with increasing temperature above 25°C, complexes of decreasing internal lipid-to-protein molar ratio are formed. However, by electron microscopy we show that the overall size of these complexes remains approximately the same, i.e., bars with dimensions $\text{70 × 220}\text{A}\text{?}$. A temperature-reversible transformation occurs between these complexes, which cannot be isolated by gel chromatography. In contrast, the complex of molar ratio 70/1 remains stable at lower temperatures.     

A study of the nature of the immediate precursor of the extracellular α-amylase of Bacillus amyloliquefaciens. A reappraisal

1. A defined medium was devised for use in washed-cell experiments with post-exponential-phase cultures of Bacillus amyloliquefaciens. The medium allowed alpha-amylase to be secreted, bacterial concentration to increase and l-[U-(14)C]valine to be incorporated into protein at a linear rate, which was the same as in a post-exponential-phase culture, for up to 6h. 2. Determination of the specific radioactivity of l-[U-(14)C]valine in the medium, the intracellular amino acid pool, the cellular protein and the isolated alpha-amylase, after a 3h incubation of washed cells in the defined medium, showed that at least 76% of the alpha-amylase secreted was synthesized de novo. 3. By isolating the alpha-amylase formed during a 6h incubation in the presence of l-[U-(14)C]valine it was shown that the specific radioactivity of the N-terminal valine, within the limits of experimental error, was the same as that of the total valine residues from the complete alpha-amylase molecule. 4. A consideration of these results in relation to the whole literature on the subject strongly supports the idea that there is no reason to suppose that extracellular alpha-amylase is formed from a high-molecular-weight precursor in B. amyloliquefaciens and closely related organisms with identical characteristics of exoenzyme secretion.