Development of doubled haploids from an elite <Emphasis Type="Italic">indica</Emphasis> rice hybrid (BS6444G) using anther culture

A total of 200 doubled haploids (DHs) were generated from an elite rice hybrid, ‘BS6444G’ for which an androgenic method was developed by manipulating the physical and chemical factors. The spike pretreated at 10?°C for 7–8 days was effective for callusing and green plant regeneration. The maximum callus frequency was achieved when the anthers cultured in N6 medium supplemented with 2.0 mg L^?1 2,4-diclorophenoxyacetic acid, 0.5 mg L^?1 6-benzylaminopurine and 3% maltose. Calli induced in N6 media also showed significant green shoot regeneration in MS medium supplemented with 0.5 mg L^?1 1-napthalene acetic acid, 0.5 mg L^?1 kinetin, 1.5 mg L^?1 benzylaminopurine and 3% sucrose producing 210 green plants. Assessment of the ploidy status showed 95.71% fertile diploids and 4.2% polyploids; no haploids were observed. A total of 38 sequence-tagged microsatellite (STMS) markers proved able to discriminate a heterozygote from all the 200 DHs. The DHs grown in the field showed significant variation for their agronomic traits. Comparison of traits with control indicates homogeneity within each DH line and significant variance of traits between DH lines. Nine DH lines produce higher grain yield than the hybrid parent which suggests the possibility of exploiting hybrid vigor in indica rice through the development of DH lines of high yielding hybrids.     

Niche-based assessment of contributions of legumes to the nitrogen economy of Western Kenya smallholder farms

Nitrogen (N) deficiency is a major constraint to the productivity of the African smallholder farming systems. Grain, green manure and forage legumes have the potential to improve the soil N fertility of smallholder farming systems through biological N₂-fixation. The N₂-fixation of bean (Phaseolus vulgaris), soyabean (Glycine max), groundnut (Arachis hypogaea), Lima bean (Phaseolus lunatus), lablab (Lablab purpureus), velvet bean (Mucuna pruriens), crotalaria (Crotalaria ochroleuca), jackbean (Canavalia ensiformis), desmodium (Desmodium uncinatum), stylo (Stylosanthes guianensis) and siratro (Macroptilium atropurpureum) was assessed using the ¹⁵N natural abundance method. The experiments were conducted at three sites in western Kenya, selected on an agro-ecological zone (AEZ) gradient defined by rainfall. On a relative scale, Museno represents high potential AEZ 1, Majengo medium potential AEZ 2 and Ndori low potential AEZ 3. Rainfall in the year of experimentation was highest in AEZ 2, followed by AEZ 1 and AEZ 3. Experimental fields were classified into high, medium and low fertility classes, to assess the influence of soil fertility on N₂-fixation performance. The legumes were planted with triple super phosphate (TSP) at 30 kg P ha^?1, with an extra soyabean plot planted without TSP (soyabean-P), to assess response to P, and no artificial inoculation was done. Legume grain yield, shoot N accumulation, %N derived from N₂-fixation, N₂-fixation and net N inputs differed significantly (P<0.01) with rainfall and soil fertility. Mean grain yield ranged from 0.86 Mg ha^?1, in AEZ 2, to 0.30 Mg ha^?1, in AEZ 3, and from 0.78 Mg ha^?1, in the high fertility field, to 0.48 Mg ha^?1, in the low fertility field. Shoot N accumulation ranged from a maximum of 486 kg N ha^?1 in AEZ 2, to a minimum of 10 kg N ha^?1 in AEZ 3. Based on shoot biomass estimates, the species fixed 25–90% of their N requirements in AEZ 2, 23–90% in AEZ 1, and 7–77% in AEZ 3. Mean N₂-fixation by green manure legumes ranged from 319 kg ha^?1 (velvet bean) in AEZ 2 to 29 kg ha^?1 (jackbean) in AEZ 3. For the forage legumes, mean N₂-fixation ranged from 97 kg N ha^?1 for desmodium in AEZ 2 to 39 kg N ha^?1 for siratro in AEZ 3, while for the grain legumes, the range was from 172 kg N ha^?1 for lablab in AEZ 1 to 3 kg N ha^?1 for soyabean-P in AEZ 3. Lablab and groundnut showed consistently greater N₂-fixation and net N inputs across agro-ecological and soil fertility gradients. The use of maize as reference crop resulted in lower N₂-fixation values than when broad-leaved weed plants were used. The results demonstrate differential contributions of the green manure, forage and grain legume species to soil fertility improvement in different biophysical niches in smallholder farming systems and suggest that appropriate selection is needed to match species with the niches and farmers’ needs.     

Bracken fern (Pteridium aquilinum L. kuhn) promotes an open nitrogen cycle in heathland soils

Background and Aims

In spite of the broad array of studies conducted on the ecology of bracken fern (Pteridium aquilinum (L.) kuhn), there is currently only a limited understanding of how P. aquilinum alters the soil environment in which it succeeds. P. aquilinum is one of the world’s most aggressive invasive species and is known to effectively invade conservation priority habitats such as Calluna vulgaris (L.) heathland. The aim of this study was to evaluate differences in soil properties between intact stands of C. vulgaris and neighboring P. aquilinum to assess how P. aquilinum alters soil N transformations in a manner that might promote its success.

Methods

Replicate plots in five independently paired stands of P. aquilinum and C. vulgaris were established on land in which P. aquilinum is actively invading. Soils under the two plant types were evaluated for total N, mineralisable N, net nitrification, nitrifier activity, denitrification enzyme activity, polyphenol N complexing capacity, and resin sorption of inorganic N.

Results

Soils under P. aquilinum were consistently higher in NO₃ ^- and NH₄ ⁺ concentrations compared to C. vulgaris. Extractable organic and inorganic N concentrations for soil under P. aquilinum were respectively 65 %, 77 % and 358 % greater in amino N NH₄ ⁺-N and NO₃ ^--N compared to that under C. vulgaris. In-situ net nitrification (NO₃ ^- sorption to ionic resins) was found to be nearly 300 times greater under P. aquilinum than under C. vulgaris.

Conclusions

P. aquilinum alters the soil environment as to create an inorganic N-rich environment that is favorable to its growth and development.     

Factors modulating cottongrass seedling growth stimulation to enhanced nitrogen and carbon dioxide: compensatory tradeoffs in leaf dynamics and allocation to meet potassium-limited growth

Eriophorum vaginatum is a characteristic species of northern peatlands and a keystone plant for cutover bog restoration. Understanding the factors affecting E. vaginatum seedling establishment (i.e. growth dynamics and allocation) under global change has practical implications for the management of abandoned mined bogs and restoration of their C-sequestration function. We studied the responses of leaf dynamics, above- and belowground biomass production of establishing seedlings to elevated CO₂ and N. We hypothesised that nutrient factors such as limitation shifts or dilutions would modulate growth stimulation. Elevated CO₂ did not affect biomass, but increased the number of young leaves in spring (+400 %), and the plant vitality (i.e. number of green leaves/total number of leaves) (+3 %), both of which were negatively correlated to [K⁺] in surface porewater, suggesting a K-limited production of young leaves. Nutrient ratios in green leaves indicated either N and K co-limitation or K limitation. N addition enhanced the number of tillers (+38 %), green leaves (+18 %), aboveground and belowground biomass (+99, +61 %), leaf mass-to-length ratio (+28 %), and reduced the leaf turnover (?32 %). N addition enhanced N availability and decreased [K⁺] in spring surface porewater. Increased tiller and leaf production in July were associated with a doubling in [K⁺] in surface porewater suggesting that under enhanced N production is K driven. Both experiments illustrate the importance of tradeoffs in E. vaginatum growth between: (1) producing tillers and generating new leaves, (2) maintaining adult leaves and initiating new ones, and (3) investing in basal parts (corms) for storage or in root growth for greater K uptake. The K concentration in surface porewater is thus the single most important factor controlling the growth of E. vaginatum seedlings in the regeneration of selected cutover bogs.     

Protein A immunosensor for the detection of immunoglobulin G by impedance spectroscopy

A novel highly sensitive electrochemical impedimetric Protein A immunosensor for the determination of immunoglobulin G (IgG) was developed by immobilization of Protein A within a newly synthesized, and characterized polymer, poly(maleicanhydride-alt-decene-1). TiO₂ nanoparticles (10–30 nm) were synthesized, characterized with X-ray diffraction, transmission electron microscopy and Brunauer–Emmett–Teller surface analysis. The electron transfer between IgG and the poly(maleicanhydride-alt-decene-1)-TiO₂-Protein A is quasireversible with a formal potential of 225 mV vs Ag|AgCl. The response of the poly(maleicanhydride-alt-decene-1)-TiO₂-Protein A immunosensor was proportional to IgG concentration with a correlation coefficient of 0.9963. The detection limit and linear range was 0.57 ng mL^?1 and 0.0062–500 μg mL^?1, respectively. Impedance measurments showed that synthesized TiO₂ nanoparticles have better conducting properties compared with commercial degussa P25 TiO₂ nanoparticles. The nonspecific binding of anti-MBP was 10 %. The label-free impedimetric immunosensor provided a simple and sensitive detection method for the specific determination of IgG in human serum.     

Binding of benzo[a]pyrene at the 1,3,6 positions to nucleic acids in vivo on mouse skin and in vitro with rat liver microsomes and nuclei

Loss of tritium from specific positions in [³H,¹⁴C] aromatic hydrocarbons can elucidate their binding site(s) to DNA and RNA and indicate the mechanism of activation. Studies of tritium loss from [6-³H,¹⁴C]benzo[a]pyrene(B[a]P), [1,3-³H,¹⁴C]B[a]P, [1,3,6-³H,¹⁴C]B[a]P, [6,7-³H,¹⁴C]B[a]P, and [7-³H,¹⁴C]B[a]P were conducted in vitro using liver nuclei and microsomes from 3-methylcholanthrene-induced Sprague-Dawley rats and in vivo on the skin of Charles River CD-1 mice. The relative loss of tritium from $\text{[}{}^3\text{H}\text{,}{}^{14}\text{C}\text{]}\text{B}\text{[a]}\text{P}$ was measured after binding to skin DNA and RNA, to nuclear DNA, and to native and denatured calf thymus and rat liver DNA's and poly(G) by microsomal activation. In skin, nuclei, and microsomes plus native DNA, virtually all B[a]P binding occurred at positions 1,3 and 6; while with microsomes plus denatured DNA or poly(G), B[a]P showed no binding at the 6 position and a small amount at the 1 and 3 positions. In vivo and with nuclei, binding at the 6 position predominated. Little loss of tritium from the 7 position was seen; this was expected because binding at this position is not thought to occur. This confirms the interpretation of loss of tritium as an indication of binding at a given position. These results demonstrate that the use of microsomes to activate B[a]P is not a valid model system for delineating the in vivo mechanism of B[a]P activation, and support previous evidence for one-electron oxidation as the mechanism of activation of hydrocarbons in binding to nucleic acids.     

The interaction of carbon-14-labelled alkyl carbamates, labelled in the alkyl and carbonyl positions, with DNA in vivo

The binding of ethyl carbamate labelled with carbon-14 in the alkyl or carbonyl group, and of methyl, n-butyl and n-propyl carbamates labelled in the alkyl group, to the DNA of mouse liver, lung and kidney has been studied in male Crackenbush mice. Only ethyl carbamate bound to liver and kidney DNA to any significant extent.The binding of ethyl carbamate labelled with carbon-14 in the C₁, C₂ or the carbonyl position was examined and compared. The levels of binding of [1-¹⁴C]- and [2-¹⁴C]ethyl carbamate to liver DNA were not significantly different (328 ± 34 and 267 ± 24 dpm/mg DNA, respectively), but there was very little binding of the [carbonyl-¹⁴C]ethyl carbamate (26 ± 3 dpm/mg DNA). Furthermore, only 18% of the radioactivity was removed from the DNA labelled with the alkyl-labelled carbamates, whereas 65% of the radioactivity was removed from the DNA labelled with carbonyl-labelled ethyl carbamate on continuous ether extraction. It was concluded that the bound molecule does not contain the carbonyl carbon and is probably an ethyl group.     

N2-Fixation and Seedling Growth Promotion of Lodgepole Pine by Endophytic Paenibacillus polymyxa

We inoculated lodgepole pine (Pinus contorta var. latifolia (Dougl.) Engelm.) with Paenibacillus polymyxa P2b-2R, a diazotrophic bacterium previously isolated from internal stem tissue of a naturally regenerating pine seedling to evaluate biological nitrogen fixation and seedling growth promotion by this microorganism. Seedlings generated from pine seed inoculated with strain P2b-2R were grown for up to 13 months in a N-limited soil mix containing 0.7 mM available N labeled as Ca(¹⁵NO₃)₂ to facilitate detection of N₂-fixation. Strain P2b-2R developed a persistent endophytic population comprising 10²–10⁶?cfu?g^?1 plant tissue inside pine roots, stems, and needles during the experiment. At the end of the growth period, P2b-2R had reduced seedling mortality by 14 % and ¹⁵N foliar N abundance 79 % and doubled foliar N concentration and seedling biomass compared to controls. Our results suggest that N₂-fixation by P. polymyxa enhanced growth of pine seedlings and support the hypothesis that plant-associated diazotrophs capable of endophytic colonization can satisfy a significant proportion of the N required by tree seedlings growing under N-limited conditions.     

Comparative studies on phosphate utilization of two bloom-forming Microcystis spp. (cyanobacteria) isolated from Lake Taihu (China)

Microcystis is a common freshwater bloom-dominating cyanobacterial genus. However, the crucial factors that affect the seasonal succession and extent of dominance of different species remain largely unknown. This study investigated inorganic phosphorus (P) uptake, growth, and P utilization of two dominant Microcystis species from Lake Taihu (China) at different P concentrations and temperatures. Compared with Microcystis wesenbergii, Microcystis flos-aquae had higher maximum uptake rate and luxury storage coefficient. However, M. flos-aquae also had a higher P demand for growth. Under P-rich conditions (64.5 to 174.5 μM), M. flos-aquae had a higher growth rate (0.16 to 0.21 d^?1) than M. wesenbergii (0.06 to 0.19 d^?1) at 22 to 30 °C. M. wesenbergii, with higher phosphate affinity, had a lower P demand to sustain its growth, and yielded a higher growth rate of 0.10 d^?1 at low P concentration (6.5 μM) at 34 °C, whereas M. flos-aquae had a lower growth rate of 0.03 d^?1. Therefore, M. flos-aquae was dominant in late spring under P-rich conditions, whereas M. wesenbergii prevailed in hot summer after M. flos-aquae decreased under P-limited conditions. These results agree well with their succession patterns in the field. Overall, succession and dominance of M. flos-aquae and M. wesenbergii in Lake Taihu are regulated by P concentration and water temperature.     

N2 fixing alder (Alnus viridis spp. fruticosa) effects on soil properties across a secondary successional chronosequence in interior Alaska

Green alder (Alnus viridis ssp. fruticosa) is a dominant understory shrub during secondary successional development of upland forests throughout interior Alaska, where it contributes substantially to the nitrogen (N) economy through atmospheric N₂ fixation. Across a replicated 200+ year old vegetation chronosequence, we tested the hypotheses that green alder has strong effects on soil chemical properties, and that ecosystem-level N inputs via N₂ fixation decrease with secondary successional stand development. Across early-, mid-, and late-successional stands, alder created islands of elevated soil N and carbon (C), depleted soil phosphorus (P), and more acidic soils. These effects translated to the stand-level in response to alder stem density. Although neither N₂ fixation nor nodule biomass differed among stand types, increases in alder densities with successional time translated to increasing N inputs. Estimates of annual N inputs by A. viridis averaged across the upland chronosequence (6.6 ± 1.2 kg N ha^?1 year^?1) are substantially less than inputs during early succession by Alnus tenuifolia growing along Alaskan floodplains. However, late-succession upland forests, where densities of A. viridis are highest, may persist for centuries, depending on fire return interval. This pattern of prolonged N inputs to late successional forests contradicts established theory predicting declines in N₂-fixation rates and N₂-fixer abundance as stands age.     

Cutting regime affects the amount and allocation of symbiotically fixed N in green manure leys

Cutting strategy effect on N₂ fixation and distribution of fixed N above and below ground in red clover (Trifolium pratense L.) and mixed red clover/perennial ryegrass (Lolium perenne L.) green manure leys was quantified in field experiments including in situ mezotrons and microplots. Symbiotically fixed N in clover, transfer of fixed N to grass in the mixed stands and the fate of ¹⁵N contained in mulch were estimated by isotope dilution. Below ground clover-derived N was estimated by leaf labelling. Total N₂ fixation was estimated by correcting fixed N in plant shoots with plant-derived N below ground and recycled N from mulch. The total N₂ fixation was larger in harvested and mulched stands (average 45 g?m^?2) than in the intact stands (32 g?m^?2). Of the fixed N, 53% (intact), 46% (harvested) and 60% (mulched) was found below ground. The average recycling of N in mulch was 21% and contributed 13.7% (pure clover) and 2.2% (mixed) of the clover N in the regrowth. Recycling of N did not decrease N₂ fixation in the mulched compared with harvested stands. The results indicate that cutting regime should be considered when estimating total amounts of N fixed by green manure leys.     

Merkel Cell Polyomavirus Small T Antigen Induces Cancer and Embryonic Merkel Cell Proliferation in a Transgenic Mouse Model

Merkel cell polyomavirus (MCV) causes the majority of human Merkel cell carcinomas (MCC) and encodes a small T (sT) antigen that transforms immortalized rodent fibroblasts in vitro. To develop a mouse model for MCV sT-induced carcinogenesis, we generated transgenic mice with a flox-stop-flox MCV sT sequence homologously recombined at the ROSA locus (ROSA ^sT), allowing Cre-mediated, conditional MCV sT expression. Standard tamoxifen (TMX) administration to adult Ubc ^CreERT2; ROSA ^sT mice, in which Cre is ubiquitously expressed, resulted in MCV sT expression in multiple organs that was uniformly lethal within 5 days. Conversely, most adult Ubc ^CreERT2; ROSA ^sT mice survived low-dose tamoxifen administration but developed ear lobe dermal hyperkeratosis and hypergranulosis. Simultaneous MCV sT expression and conditional homozygous p53 deletion generated multi-focal, poorly-differentiated, highly anaplastic tumors in the spleens and livers of mice after 60 days of TMX treatment. Mouse embryonic fibroblasts from these mice induced to express MCV sT exhibited anchorage-independent cell growth. To examine Merkel cell pathology, MCV sT expression was also induced during mid-embryogenesis in Merkel cells of Atoh1 ^CreERT2/+; ROSA ^sT mice, which lead to significantly increased Merkel cell numbers in touch domes at late embryonic ages that normalized postnatally. Tamoxifen administration to adult Atoh1 ^CreERT2/+; ROSA ^sT and Atoh1 ^CreERT2/+; ROSA ^sT; p53f ^lox/flox mice had no effects on Merkel cell numbers and did not induce tumor formation. Taken together, these results show that MCV sT stimulates progenitor Merkel cell proliferation in embryonic mice and is a bona fide viral oncoprotein that induces full cancer cell transformation in the p53-null setting.     

Nitrogen-addition effects on leaf traits and photosynthetic carbon gain of boreal forest understory shrubs

Boreal coniferous forests are characterized by fairly open canopies where understory vegetation is an important component of ecosystem C and N cycling. We used an ecophysiological approach to study the effects of N additions on uptake and partitioning of C and N in two dominant understory shrubs: deciduous Vaccinium myrtillus in a Picea abies stand and evergreen Vaccinium vitis-idaea in a Pinus sylvestris stand in northern Sweden. N was added to these stands for 16 and 8 years, respectively, at rates of 0, 12.5, and 50 kg N ha^?1 year^?1. N addition at the highest rate increased foliar N and chlorophyll concentrations in both understory species. Canopy cover of P. abies also increased, decreasing light availability and leaf mass per area of V. myrtillus. Among leaves of either shrub, foliar N content did not explain variation in light-saturated CO₂ exchange rates. Instead photosynthetic capacity varied with stomatal conductance possibly reflecting plant hydraulic properties and within-site variation in water availability. Moreover, likely due to increased shading under P. abies and due to water limitations in the sandy soil under P. sylvestris, individuals of the two shrubs did not increase their biomass or shift their allocation between above- and belowground parts in response to N additions. Altogether, our results indicate that the understory shrubs in these systems show little response to N additions in terms of photosynthetic physiology or growth and that changes in their performance are mostly associated with responses of the tree canopy.     

Effects of stationary phase elongation and initial nitrogen and phosphorus concentrations on the growth and lipid-producing potential of Chlorella sp. HQ

Higher lipid production and nutrient removal rates are the pursuing goals for synchronous biodiesel production and wastewater treatment technology. An oleaginous alga Chlorella sp. HQ was tested in five different synthetic water, and it was found to achieve the maximum biomass (0.27 g L^?1) and lipid yield (41.3 mg L^?1) in the synthetic secondary effluent. Next, the effects of the stationary phase elongation and initial nitrogen (N) and phosphorus (P) concentrations were investigated. The results show that the algal characteristics were affected apparently under different N concentrations but not P, which were verified by Logistic and Monod models. At the early stationary phase, the algal biomass, lipid and triacylglycerols (TAGs) yields, and P removal efficiency increased and reached up to 0.19 g L^?1, 46.7 mg L^?1, 14.3 mg L^?1, and 94.3 %, respectively, but N removal efficiency decreased from 86.2 to 26.8 % under different N concentrations. And the largest TAGs yield was only 6.4 mg L^?1 and N removal efficiency was above 71.1 % under different P concentrations. At the late stationary phase, the maximal biomass, lipid and TAGs yields, and P removal efficiencies primarily increased as the initial N and P concentrations increase and climbed up to 0.49 g L^?1, 99.2 mg L^?1, 54.0 mg L^?1, and 100.0 %, respectively. It is concluded that stationary phase elongation is of great importance and the optimal initial N/P ratio should be controlled between 8/1 and 20/1 to serve Chlorella sp. HQ for better biodiesel production and secondary effluent purification.     

Litter- and soil carbohydrate-carbon stocks in 2-, 4- and 10-year-old improved fallows in eastern Zambia

This study evaluated the effects of tree species and sites on soil carbohydrates, litterfall, and litter chemistry in 2-, 4- and 10-year-old improved fallows at three sites in eastern Zambia. Between April 2002 and August 2003, litter was collected in 2-year-old tree fallows at Kalichero, Kalunga and Msekera for chemical analyses. Soil samples collected at 0–30 cm from all experiments were analysed for total soil organic carbon (SOC), but only those from 4- and 10-year-old fallows were analysed for carbohydrates. Soil arabinose- and mannose-C stocks, and carbohydrate-C percentages of SOC (7.7–20.6 %) significantly (P < 0.05) differed across tree species in 10-year-old coppicing fallows at Msekera. Converting M + F to improved fallows resulted in a decline in monosaccharide-C, carbohydrate-C stocks and carbohydrate-C percentage of SOC. There were significant (P < 0.05) variations in litterfall (0.7–2.3 t ha^?1 year^?1) and litter C contents (0.3–1.1 t ha^?1 year^?1) across 2-year-old coppicing tree fallows at Msekera. Litter production and C contents were significantly greater on sandy soils at Kalunga than on fine-textured soils at Msekera. Litter chemical contents (C, N, AUR and polyphenols) and ratios (C:N, P:N, AUR:N, and (AUR + P):N) for litter in fallows differed significantly (P < 0.05) across species and sites. In this study, the role of litter in carbon cycling in improved fallows depended on tree species and site conditions.     

Heterologous overexpression of Vigna radiata epoxide hydrolase in Escherichia coli and its catalytic performance in enantioconvergent hydrolysis of p-nitrostyrene oxide into (R)-p-nitrophenyl glycol

Two native epoxide hydrolases (EHs) were previously discovered from mung bean powder (Vigna radiata), both of which can catalyze the enantioconvergent hydrolysis of p-nitrostyrene oxide (pNSO). In this study, the encoding gene of VrEH1 was successfully cloned from the cDNA of V. radiata by RT-PCR and rapid amplification of cDNA ends (RACE) technologies. High homologies were found to two putative EHs originated from Glycine max (80 %) and Medicago truncatula (79 %). The vreh1 gene constructed in pET28a(+) vector was then heterologously overexpressed in Escherichia coli BL21(DE3), and the encoded protein was purified to homogeneity by nickel affinity chromatography. It was shown that VrEH1 has an optimum activity at 45 °C and is very thermostable with an inactivation energy of 468 kJ mol^-1. The enzyme has no apparent requirement of metal ions for activity, and its activity was strongly inhibited by 1 mM of Ni²⁺, Cu²⁺, Fe²⁺, or Co²⁺. By adding 0.1 % Triton X-100, the enzyme activity could be significantly increased up to 340 %. VrEH1 shows an unusual ability of enantioconvergent catalysis for the hydrolysis of racemic pNSO, affording (R)-p-nitrophenyl glycol (pNPG). It displays opposite regioselectivity toward (S)-pNSO (83 % to C_α) in contrast to (R)-pNSO (87 % to C_β). The K _M and k _cat of VrEH1 were determined to be 1.4 mM and 0.42 s^-1 for (R)-pNSO and 5.5 mM and 6.2 s^-1 for (S)-pNSO. This thermostable recombinant VrEH1 with enantioconvergency is considered to be a promising biocatalyst for the highly productive preparation of enantiopure vicinal diols and also a good model for understanding the mechanism of EH stereoselectivity.     

The fate of 15N-nitrate in a northern peatland impacted by long term experimental nitrogen, phosphorus and potassium fertilization

Information about the impact of nitrogen (N) deposition on the fate of deposited N in peatland ecosystems is lacking. Thus we investigated the fate of experimentally added ¹⁵N in long-term N-fertilized treatments in a Sphagnum-dominated ombrotrophic bog. Fertilization significantly stimulated vascular plant and suppressed Sphagnum and Polytrichum moss growth. N content in peat, mosses, and vascular plants was raised by the fertilizer addition and reached a maximum at 3.2 g m^?2 N input level with phosphorus (P) and potassium (K) addition. Most of N was retained in the vegetation and upper 10 cm of the peat. When N deposition equalled 1.6 g m^?2 and less, or 3.2 g m^?2 N with P and K addition, no inorganic N leaching was observed on the plots. This result indicates that co-fertilization with P and K raised the N retention capacity and that critical N loads with respect to N saturation depend on P and K availability. Most of the deposited ¹⁵N was recovered in the bulk peat, which may be related to a rapid immobilization of inorganic N by microorganisms and mycorrhizal assimilation. Increase of N, P, and K fertilization increased the contribution of vascular plants to N retention significantly and reduced those of mosses. The increase was mainly related to enhanced productivity, vascular biomass and N content in tissues; the reduced retention by mosses resulted from both reduced moss biomass and assimilation. The study shows that the N filter function of ombrotrophic bogs will be influenced by interactions with other nutrients and shifts in plant community structure.     

Binding of the chemical carcinogen N-hydroxy-acetyl-aminofluorene to ploidy classes of rat liver nuclei as separated by velocity sedimentation at unit gravity

A large sedimentation device was developed that allows separation of 5 × 10⁸ rat liver nuclei by velocity sedimentation at unit gravity. Using the apparatus isolated rat liver nuclei were separated into classes of diploid stromal (Von Kuppfer, sinusoidal lining) nuclei, diploid parenchymal nuclei and tetraploid parenchymal nuclei respectively. DNA content and volume of the nuclei were measured. Diploid nuclei were 100% pure; tetraploid nuclei 98%.The in vivo binding of the liver carcinogen [³H]-N-hydroxy-AAF to these classes of nuclei was determined (total binding to protein, DNA and RNA). Binding and the subsequent removal of the fluorene derivatives was registered as a function of time. At all stages diploid stromal nuclei bound 2.6–5 times less carcinogen than did diploid parenchymal nuclei. Tetraploid parenchymal nuclei bound more than twice (2.3–3.95) the amount, that was present in their diploid counterpart. This effect became more pronounced 11 days after application of N-hydroxy-N-acetyl-2-aminofluorene.DNA was enzymatically purified from pooled classes of the various nuclear types. For purified DNA also it was found that DNA derived from diploid stromal nuclei bound 2.6–2.8 times less carcinogen than did DNA derived from diploid parenchymal nuclei.     

Nitrogen Fertilization Does Significantly Increase Yields of Stands of Miscanthus × giganteus and Panicum virgatum in Multiyear Trials in Illinois

The C4 perennial grasses Miscanthus × giganteus and Panicum virgatum (switchgrass) are emerging bioenergy crops. They are attractive because they are productive and recycle nutrients to the overwintering belowground rhizomes, before the dry shoots are harvested. They are therefore expected to require minimal fertilizer inputs, a desirable trait for any crop. Until now, Europe has had the only long-term trials of M. × giganteus, and these have either shown no or a small effect of nitrogen fertilization, while trials of P. virgatum in the USA have shown a clear positive effect of N fertilization. This study exploited the first long-term trials of M. × giganteus in the USA, and first side-by-side comparison with P. virgatum, to test the hypothesis that N fertilization would only improve yields of the latter. A split-plot N fertility treatment (0, 67, 134, and 202 kg(N)?ha^?1) was added to >5-year-old replicated stands of the two crops at seven locations on contrasting soils in the US Midwest. Averaged across all locations, M. × giganteus yields increased significantly from 23.4 Mg ha^?1 with no N fertilization to 28.9 Mg ha^?1 (+25 %) at a N application rate of 202 kg ha^?1. P. virgatum also showed significant yield increases from 10.33 Mg ha^?1 at 0 kg(N)?ha^?1 to 13.6 Mg ha^?1 (+32 %) at 202 kg(N)?ha^?1. Both species therefore responded to N fertilization and to a similar extent. The increase per unit of added N was small compared to crops such as Zea mays and unlikely to be economically worthwhile. Nitrogen fertilization arrested most of the long-term yield decline that would otherwise have occurred in P. virgatum, but eliminated only about 40 % of the decline observed in M. × giganteus, suggesting additional causal factors for long-term yield decline in this crop. While the crops responded to nitrogen addition at some locations, they did not at others. Therefore a one-case-fits-all optimum fertilization rate cannot be prescribed.