Genetic transformation of alfalfa somatic embryos and their clonal propagation through repetitive somatic embryogenesis

Genetically transformed alfalfa (Medicago sativa L., cv. Zajearska 83) plantlets were obtained by inoculating somatic embryos with Agrobacterium tumefaciens strains A281/pGA472 and LBA4404/pBI121. Single somatic embryos, 5–7 mm long, were released from a repetitively embryogenic culture, wounded, and cocultivated with the bacteria. The agar-solidified culture medium contained mineral salts, vitamins, 40 g l^–1 sucrose, 1 g l^–1 yeast extract and 0.05 mg l^–1 BA. Five clones, transformed with A281/pGA472, and 4 clones transformed with LBA4404/pBI121, were selected for proliferation by repetitive somatic embryogenesis, on media containing 100 mg l^–1 of kanamycin. The transformation of kanamycin-resistant clones was confirmed by assaying the activity of neomycin phosphotransferase II and/or -glucuronidase enzymes, and by the Southern blot analysis. It is suggested that the transformation/regeneration system based on somatic embryogenesis may be suitable for establishing transgenic alfalfa lines. The relatively low frequency of embryo transformation is compensated for by abundant proliferation in secondary somatic embryogenesis.Abbreviations BA 6-benzyladenine - GUS -glucuronidase - Km kanamycin - NPTII neomycin phosphotransferase II - X-gluc 5-bromo-4-chloro-3-indolyl--glucuronic acid - BM basal medium     

Metabolic changes during rooting in stem cuttings of five mangrove species

Vegetative propagation through rooting in stem cuttings in five tree mangroves namely Bruguiera parviflora, Cynometra iripa, Excoecaria agallocha, Heritiera fomes, and Thespesia populnea using IAA, IBA and NAA was reported. Spectacular increase in the root number was noted in the cuttings of H. fomes and C. iripa treated together with IBA (5000 ppm) and NAA (2500 ppm). The highest number of roots was obtained with IBA (2500 ppm) and NAA (500 ppm) in E. agallocha. B. parviflora and T. populnea responded better to IAA and IBA treatment. The species specific variation in the rooting response to exogenous application of auxins was reflected in the metabolic changes during initiation and development of roots in cuttings. Biochemical analysis showed increase of reducing sugar in the above-girdled tissues at initiation as well as subsequent development of roots which was further enhanced by the use of auxins. Decreases in the total sugar, total carbohydrate and polyphenols and increase in total nitrogen were recorded in the girdled tissues and the high C/N ratio at the initial stage helped in initiation of roots in all the species. Interaction of IBA and NAA promoted starch hydrolysis better than IAA and IBA during root development and subsequently reduced the C/N ratio and increased the protein-nitrogen activity during root development which suggest the auxin influenced mobilization of nitrogen to the rooting zone.Abbreviations IAA Indole-3-acetic acid - IBA Indole-butyric acid - NAA A-naphthalene acetic acid     

Gibbard's evolutionary theory of rationality and its ethical implications

Gibbard's theory of rationality is evolutionary in terms of its result as well as its underpinning argument. The result is that judgments about what is rational are analyzed as being similar to judgments of morality — in view of what Darwin suggests concerning the latter. According to the Darwinian theory, moral judgments are based on sentiments which evolve to promote the survival and welfare of human societies. On Gibbard's theory, rationality judgments should be similarly regarded as expressing emotional attachments to behavioral norms which originate and function to coordinate social interaction. Consequently, Gibbard's theory of rationality might be used to illuminate Darwin's theory of morality, and vice versa. Additionally, as argued in the present essay, both can be further elaborated, and defended, by developing related themes in philosophical ethics: viz., connected with Hume and 20th-century emotivists. The main problem is that this general Darwinian approach faces widespread opposition nowadays, not only in ethics but in philosophy of science. The purpose of this essay is to analyze Gibbard's theory, critically and constructively, with emphasis on the pertinent commonalities in Darwin, Hume and the emotivists, while also critically addressing their common enemies. The pervasive methodological orientation is to relate this analysis to (philosophy of) science in general, and biological science in particular.     

Fatty acid composition of vitellogenin from four teleost species

The fatty acid compositions of vitellogenin and liver from cod (Gadus morhua), rainbow trout (Oncorhynchus mykiss), turbot (Scophthalmus maximus) and wolffish (Anarhichas lupus) were determined. Vitellogenin was isolated from plasma of estradiol-17-treated fish by precipitation with EDTA-Mg²⁺ and distilled water or by high-performance ion-exchange chromatography. In all investigated species, vitellogenin contained 16–18% (w/w) lipid, in which polyunsaturated fatty acids, predominantly 20:5 (n-3) and 22:6 (n-3), comprised about 50% of the total fatty acids. The proportions of saturated, monounsaturated, polyunsaturated and (n-3) fatty acids in vitellogenin of the different species were generally similar, although the relative content of specific fatty acids was distinctive for each species. The distribution of fatty acids in total lipids of vitellogenin was highly consistent among individual females of each species. In contrast, liver fatty acid composition varied considerably, both within and between species. Altogether, the differences in the fatty acid composition of vitellogenin and liver from each species indicate that a specific selection of fatty acids occurs during the lipidation of vitellogenin.Abbreviations BHT butylated hydroxytoluene - E-17 estradiol-17 - EDTA ethylenedinitrilo tetra-acetic acid disodium salt dihydrate - FA fatty acids - FAME fatty acid methyl esters - HDL high density lipoproteins - PUFA polyunsaturated fatty acids - SD standard deviation - TLC thin-layer chromatography - VHDL very high density lipoproteins - VLDL very low density lipoproteins - v/v volume per volume - w/v weight per volume - w/w weight per weight     

In vitro investigation of α-amylase release from the digestive cells of the bivalve mollusc Pecten maximus: effect of second messengers and biogenic amines

The (neuro)endocrine control of enzyme release from invertebrate digestive cells remains poorly understood. A tissue dissociation procedure was developed to investigate the regulatory mechanisms of -amylase discharge from the cells of the stomach-digestive gland complex of the scallop Pecten maximus. The validity of the experimental system was tested by increasing the intracellular concentration of second messenger analogues (N ⁶,2-o-dibutyryl-adenosine-3,5 cyclic monophosphate and the ionophore A23187) known to mimic the activity of naturally occurring secretagogues in vertebrates: N ⁶,2-o-dibutyryl-adenosine-3,5 cyclic monophosphate increased the time and dose-dependent release of -amylase in a similar way as in vertebrates. A23187 was also very effective in inducing enzyme discharge. Since the in vitro bioassay was shown to be functional and because axon terminals were previously seen in close contact to -amylase secreting cells, the effect of some classic neurotransmitters was explored. Only the cholinergic agonist carbachol and dopamine evoked a secretory response. Maximal stimulation of -amylase release was reached at 10^-5 mol·l^-1 carbachol; at the same concentration dopamine was less effective than carbachol. By contrast, serotonin was totally inactive. The in vitro bioassay should prove useful for the identification of regulatory molecules involved in the control of enzyme discharge and to study stimulus secretion coupling mechanisms in scallop digestive cells.Abbreviations DBcAMP N ⁶, 2-O-dibutyryl-adenosine-3,5 cyclic monophosphate - cAMP adenosine-3,5 cyclic monophosphate     

Frequency-dependent interference by magnetic fields of nerve growth factor-induced neurite outgrowth in PC-12 cells

We have shown that 50 Hz sinusoidal magnetic fields within the 5-10 micro Tesla (μT) rms range cause an intensity-dependent reduction in nerve growth factor (NGF) stimulation of neurite outgrowth (NO) in PC-12 cells. Here we report on the frequency dependence of this response over the 15-70 Hz range at 5 Hz intervals. Primed PC-12 cells were plated in collagen-coated, 60 mm plastic petri dishes with or without 5 ng/ml NGF and were exposed to sinusoidal magnetic fields for 22 h in a CO₂ incubator at 37 °C. One 1,000-turn coil, 20 cm in diameter, generated vertically oriented magnetic fields. The dishes were stacked on the center axis of the coil to provide a range of intensities between 3.5 and 9.0 μT rms. The flux density of the ambient DC magnetic field was 37 μT vertical and 19 μT horizontal. The assay consisted of counting over 100 cells in the central portion (radius ≤0.3 cm) of each dish and scoring cells positive for NO. Sham exposure of cells treated identically with NGF demonstrated no difference in the percentage of cells with NO between exposed and magnetically shielded locations within the incubator. Analysis of variance demonstrated flux density-dependent reductions in NGF-stimulated NO over the 35-70 Hz frequency range, whereas frequencies between 15 Hz and 30 Hz produced no obvious reduction. The results also demonstrated a relative maximal sensitivity of cells at 40 Hz with a possible additional sensitivity region at or above 70 Hz. These findings suggest a biological influence of perpendicular AC/DC magnetic fields different from those identified by the ion parametric resonance model, which uses strictly parallel AC/DC fields. © 1995 Wiley-Liss, Inc.     

Food dependent color patterns inThamnocephalus platyurus Packard (Branchiopoda: Anostraca); a laboratory study

Coloration of phyllopods varies from place to place and from one life stage to another. It ranges from translucent or whitish through gray, blue, green, orange, and reddish. Here, we present experimental evidence for a food- dependent color pattern inThamnocephalus platyurus Packard. The presence or absence of the synthetic pigment trans — — carotene in a baker's yeast diet was the controlling factor. All the 24 old larvae used in the experiment were whitish in color. From day 6 until the end the experiment (day 11), 100% of the shrimps under a diet with synthetic trans — — carotene (treatment 1) exhibited a characteristic color pattern which consisted of an orange color in the cercopods, and in all theracopods; the rest of the body exhibited no particular color. In comparison, 100% of the shrimps under a diet without synthetic trans — — carotene (treatment 2) were whitish throughout the body. In females from treatment 1, the ovaries and oocytes were green-bluish, while in females from treatment 2 the ovaries and oocytes were whitish. No significant differences in survival and growth were found, except that at day 9, there was a significant difference in growth, the females with the synthetic trans — — carotene group growing faster.     

Radiative properties of hardwood leaves to ultraviolet irradiation

Spectral reflectance and transmittance of leaves to ultraviolet irradiation were determined under laboratory conditions for seven species of hardwood trees, namely red oak (Quercus rubra, L), black oak (Q. velutina, Lamarch), white oak (Q. alba, L.), sugar maple (Acer saccharum), Norway maple (A. plantanoides), hickory (Carya tomemtosa), sweetgum (Liquidambar styraciflua), and black oak litter. The experimental system consisted of a solar simulator, an integrating sphere, and a spectroradiometer. Experiments were repeated three to five times for both adaxial and abaxial surfaces of fresh leaves chosen at randomly. The spectral distributions and simple averages of the radiative properties in the wavelength ranges of ultraviolet-B (UV-B, 280–320 nm) and ultraviolet-A (UV-A, 320–400 nm) were determined. The spectral distributions of reflectance were similar between adaxial and abaxial surfaces, although the magnitude varied among tree species. Leaf reflectance was very low for the ultraviolet spectrum in general and varied among species and between adaxial and abaxial surfaces. It was generally higher over the UV-A waveband compared to UV-B, and higher on the abaxial than adaxial surface. The broadband reflectance in the UV-A range (over all species) was 5.0 and 3.9% for abaxial and adaxial surface, respectively, compared to 3.5 and 2.8% in UV-B. The transmittance through leaves was extremely small in the UV-B (<0.1%) and nearly zero in the UV-A spectral range. Consequently, the absorptance of ultraviolet radiation by leaves, as determined from the measured reflectance and transmittance, was quite high, being more than 90% for all the combinations of species and wavebands examined. The reported results are useful for studies requiring spectral radiative properties of the examined leaves with respect to ultraviolet irradiation.     

Variation in growth and age at maturity in bluegill sunfish: genetic or environmental effects?

Populations of bluegill sunfish Lepomis macrochirus , experiencing heavy juvenile predation, showed increased growth rates and increased age and size at maturity relative to populations experiencing decreased predation on juveniles but increased predation on adults. This study examined bluegills experimentally from both types of populations and a cross between them in a common environment to determine if variation in growth and age at maturity is genetically or environmentally induced. Two factorial experiments, varying strain of bluegills and resource availability, were used to evaluate differences in growth rate. One experiment, varying strain of bluegills, was used to assess differences in age at maturity. Growth was strongly influenced by resource level, but growth rate did not vary among populations. Nearly all bluegills in each population matured at 1 year of age in a common environment. Thus, variation observed in source populations must be mostly attributable to differences in the environment between populations. At least three factors could potentially cause differences in growth and age at maturity: (1) variation in resource availability; (2) variation in demographic structure; and (3) variation in size-specific mortality rates caused by differences in predator abundance between populations. Observed patterns of variation between populations are best explained by effects of differences in predator populations.