Characterization of two class II chitinase genes from peanut and expression studies in transgenic tobacco plants

Two different genes encoding class II chitinases from peanut (Arachis hypogaea L. cv. NC4), A.h.Chi2;1 and A.h.Chi2;2, have been cloned. In peanut cell suspension cultures, mRNA levels of A.h.Chi2;2 increased after ethylene or salicylate treatment and in the presence of conidia from Botrytis cinerea. The second gene, A.h.Chi2;1, was only expressed after treatment with the fungal spores. Transgenic tobacco plants containing the complete peanut A.h.Chi2;1 gene exhibited essentially the same expression pattern in leaves as observed in peanut cell cultures. Expression characteristics of transgenic tobacco carrying a promoter-GUS fusion of A.h.Chi2;1 are described.     

Physiological aspects of Taxus brevifolia seeds in relation to seed storage characteristics

Water relations, desiccation tolerance and longevity of Taxus brevifolia (Nutt.) seeds were studied to determine the optimal stage of development and storage conditions for seeds of this species. Seeds equilibrated to a range of relative humidities (RHs) had unusually low water contents which can be accounted for by the high lipid content of gametophyte tissues (71% of the dry mass). Water relations of embryonic tissue were more typical of those reported for other seed species. The water content below which freezing transitions were not observable in the embryo was ca 0.24 g H₂O (g dry weight)⁻¹ (g g⁻¹) for all maturity classes studied. Embryos did not achieve significant levels of desiccation tolerance (survival to water contents less than 0.5 g g⁻¹) until the latter stages of development when dry matter was maximal. Mature embryos could be dried to 0.025 g g⁻¹ (seed water content of 0.010 g g⁻¹) with no loss of viability. Thus, at the latter stages of development, embryo water content could be optimized to avoid both desiccation and freezing damage. Survival of mature seeds declined over a 2-year period when seeds were stored at temperatures between 5 and 35°C and RHs between 14 and 75%, corresponding to seed water contents between 0.015 and 0.07 g g⁻¹. The deterioration rate was slowest for seeds stored at the lowest RH and temperature. Our data indicate that seeds of Taxus brevifolia show orthodox rather than recalcitrant storage characteristics, but that the optimum water content for storage was extremely low. The results suggest that even if stored at optimal water contents and low temperatures, T. brevifolia seeds will be relatively short lived. The high quantity of lipids or reducing sugars may be contributing factors in the poor storage characteristics.     

Cold acclimation and photoinhibition of photosynthesis in Scots pine

Cold acclimation of Scots pine did not affect the susceptibility of photosynthesis to photoinhibition. Cold acclimation did however cause a suppression of the rate of CO₂ uptake, and at given light and temperature conditions a larger fraction of the photosystem II reaction centres were closed in cold-acclimated than in nonacclimated pine. Therefore, when assayed at the level of photosystem II reaction centres, i.e. in relation to the degree of photosystem closure, cold acclimation caused a significant increase in resistance to photoinhibition; at given levels of photosystem II closure the resistance to photoinhibition was higher after cold acclimation. This was particularly evident in measurements at 20° C. The amounts and activities of the majority of analyzed active oxygen scavengers were higher after cold acclimation. We suggest that this increase in protective enzymes and compounds, particularly Superoxide dismutase, ascorbate peroxidase, glutathione reductase and ascorbate of the chloroplasts, enables Scots pine to avoid excessive photoinhibition of photosynthesis despite partial suppression of photosynthesis upon cold acclimation. An increased capacity for light-induced de-epoxidation of violaxanthin to zeaxanthin upon cold acclimation may also be of significance.Abbreviations APX ascorbate peroxidase - DHA dehydroascorbate - DHAR dehydroascorbate reductase - Fm maximal fluorescence when all reaction centres are closed - Fv/Fm maximum photochemical yield of PSII - GR glutathione reductase - GSH reduced glutathione - Je rate of photosynthetic electron transport - MDAR monodehydroascorbate reductase - q_N nonphotochemical quenching of fluorescence - q_P photochemical quenching of fluorescence - SOD superoxide dismutase This work was supported by the Swedish Natural Science Research Council and the National Natural Science Foundation of China.     

DPB1 * BR: an Mhc class II DPB1 allele (DPB1 * 6601) of negroid origin

The nucleotide sequence data reported in this paper have been submitted to the EMBL nucleotide sequence database and have been assigned the accession number X96986. The nameDPB1 ^* 6601 was officially assigned by the WHO Nomenclature Committee in May 1996. This follows the agreed policy that, subject to the conditions stated in the most recent Nomenclature Report (Bodmer et al. 1995), names will be assigned to new sequences as they are identified. Lists of such sequences will be published in the following WHO Nomenclature Report     

Purified moniliformin does not affect the force or rate of contraction of isolated guinea pig atria

Aspergillus flavus Link ex Fries and A. parasiticus Speare can invade peanut kernels and under certain environmental conditions produce unacceptable levels of the mycotoxin aflatoxin. A concerted effort is underway to reduce aflatoxin contamination in peanut and peanut products. A potentially effective method of control in peanut is the discovery and use of genes for resistance to either fungal invasion or aflatoxin formation. The objective of the present experimental study was to develop an effective and efficient procedure for screening individual plants or pods of single plants for resistance to invasion by the aflatoxigenic fungi and subsequent aflatoxin production. Methods of obtaining adequate drought-stress and fungal infection were developed through this series of experiments. By completely isolating the pods from the root zone and imposing drought-stress only on pegs and pods, high levels of fungal infection were observed. High amounts of preharvest aflatoxin accumulation were also produced by completely isolating the pods from the root zone. Mid-bloom inoculation with A. parasiticus-contaminated cracked corn and drought-stress periods of 40 to 60 days were the most effective procedures. This technique was used to assess peanut genotypes previously identified as being partially resistant to A. parasiticus infection or aflatoxin contamination, and segregating populations from four crosses. Variability in aflatoxin contamination was found among the 11 genotypes evaluated, however, none were significantly lower than the standard cultivars. Broad-sense heritability of four crosses was estimated through evaluation of seed from individual plants in the F₂ generation. The heritability estimates of crosses GFA-2 × NC-V11 and Tifton-8 × NC-V11 were 0.46 and 0.29, respectively, but mean aflatoxin contamination levels were high (73,295 and 27,305 ppb). This greenhouse screening method could be an effective tool when genes for superior aflatoxin resistance are identified.Cooperative investigation of the USDA-ARS and the University of Georgia, College of Agriculture.     

Development of an L6 myoblast in vitro model of moniliformin toxicosis

L6 myoblasts were used as an in vitro model to investigate the role of moniliformin and its interaction with monensin in turkey knockdown syndrome and sudden death syndromes in poultry. Cell viability and microscopic and ultrastructural alterations noted in L6 myoblasts cultured in the presence of moniliformin (0.0–0.3 g/l) were compared to those observed in parallel cultures also containing one of the following compounds: selenium (0–0.004 ng/l), thiamine (0–0.3 g/l), or pyruvate (0–0.46 g/l). Marked dilation of the RER, membranous whorls, glycogen deposition, membrane-bound cytoplasmic inclusions and necrosis were observed in myoblasts exposed to 0.03/2-0.30 g moniliformin/l medium. Supplementation of medium with thiamine and pyruvate, or selenium, provided significant protection to cells exposed to 0.0–0.3 g/l or 0.0–0.15 g moniliformin/l, respectively. Dose-dependent differences in protein and ATP production were not detected. Myoblasts grown in medium containing 0–0.15 g moniliformin/l and 7.5–50.0 M A23187, beauvericin or monensin had degrees of cytotoxicity similar to parallel cultures receiving only an ionophore. L6 myoblasts were a useful model of moniliformin toxicosis. The findings of this study suggest cytotoxicity due to moniliformin in L6 myoblasts may be due in part to oxidative damage and altered pyruvate metabolism, and that moniliformin does not predispose myoblasts to ionophore toxicosis. This study supports the results of in vivo investigations in poultry that moniliformin and monensin do not act synergistically to induce knockdown or monensin toxicosis.     

Theoretical Basis of Protocols for Seed Storage III. Optimum Moisture Contents for Pea Seeds Stored at Different Temperatures

In previous work, we demonstrated that there was an optimummoisture level for seed storage at a given temperature (Vertucciand Roos, 1990), and suggested, using thermodynamic considerations,that the optimum moisture content increased as the storage temperaturedecreased (Vertucci and Roos, 1993b). In this paper, we presentdata from a two year study of aging rates in pea (Pisum sativum)seeds supporting the hypothesis that the optimum moisture contentfor storage varies with temperature. Seed viability and vigourwere monitored during storage under dark or lighted conditionsat relative humidities between 1 and 90%, and temperatures between-5 and 65°C. The optimum moisture content varied from 0·015g H₂O g^-1 d.wt at 65°C to 0·101 g H₂O g^-1 d.wt at15°C under dark conditions and from 0·057 at 35°Cto 0·092 g H₂O g^-1 d.wt at -5°C under lighted conditions.Our results suggest that optimum moisture contents cannot beconsidered independently of temperature. This conclusion hasimportant implications for 'ultra-dry' and cryopreservationtechnologies.Copyright 1994, 1999 Academic Press Seed storage, seed aging, seed longevity, water content, temperature, glass, desiccation damage, ultradry, Pisum sativum L., pea, cryopreservation     

Saturated Molecular Map of the Rice Genome Based on an Interspecific Backcross Population

A molecular map has been constructed for the rice genome comprised of 726 markers (mainly restriction fragment length polymorphisms; RFLPs). The mapping population was derived from a backcross between cultivated rice, Oryza sativa, and its wild African relative, Oryza longistaminata. The very high level of polymorphism between these species, combined with the use of polymerase chain reaction-amplified cDNA libraries, contributed to mapping efficiency. A subset of the probes used in this study was previously used to construct an RFLP map derived from an inter subspecific cross, providing a basis for comparison of the two maps and of the relative mapping efficiencies in the two crosses. In addition to the previously described PstI genomic rice library, three cDNA libraries from rice (Oryza), oat (Avena) and barley (Hordeum) were used in this mapping project. Levels of polymorphism detected by each and the frequency of identifying heterologous sequences for use in rice mapping are discussed. Though strong reproductive barriers isolate O. sativa from O. longistaminata, the percentage of markers showing distorted segregation in this backcross population was not significantly different than that observed in an intraspecific F(2) population previously used for mapping. The map contains 1491 cM with an average interval size of 4.0 cM on the framework map, and 2.0 cM overall. A total of 238 markers from the previously described PstI genomic rice library, 250 markers from a cDNA library of rice (Oryza), 112 cDNA markers from oat (Avena), and 20 cDNA markers from a barley (Hordeum) library, two genomic clones from maize (Zea), 11 microsatellite markers, three telomere markers, eleven isozymes, 26 cloned genes, six RAPD, and 47 mutant phenotypes were used in this mapping project. Applications of a molecular map for plant improvement are discussed.     

Evidence that Prostaglandin E2 Can Block Calcium-Activated 86Rb Efflux from Rat Brain Synaptosomes via a Protein Kinase C-Dependent Mechanism

Abstract: The effects of prostaglandin E₂ (PGE₂) on ⁸⁶Rb efflux from rat brain synaptosomes were studied to explore its role in nerve ending potassium (K⁺) channel modulation. A selective dose-dependent inhibition of the calcium-activated charybdotoxin-sensitive component of efflux was found upon application of PGE₂. No significant effect was seen on basal and voltage-dependent components over the concentration range of 10–⁸ to 10–⁵M. The protein kinase C (PKC) inhibitors H-7 (10 μM) and staurosporine (100 nM), as well as prolonged preincubation (90 min) with 40-phorbol 12, 13-dibutyrate, which has been reported to down-regulate PKC, abolished the PGE₂-in- duced inhibition, whereas HA1004 (10 μM) and Rp-3′,5’cyclic phosphorothioate (100 nM), which are relatively more selective for protein kinase A than PKC, did not. 4β-Phorbol 12, 13-dibutyrate (100 nM), an activator of PKC, produced a similar inhibition of the Ca²⁺-dependent component of ⁸⁶Rb efflux but also had no effect on the basal and voltage-dependent components. These data suggest that PGE₂ can inhibit rat brain nerve ending calcium-activated ⁸⁶Rb efflux, and this inhibition may involve PKC activation.