A general method for isolation of Mu d1-8(Aprlac) operon fusions in Salmonella typhimurium LT2 from Tn10 insertion strains: chlC::Mu d1-8

Summary A general method was developed for the isolation of Salmonella thyphimurium LT2 Mu d1–8 (Ap^r lac) operon fusions in a gene displacing a Tn10 insertion. Random Mu d1–8 fusion pools were prepared to grow phage P22 lysates which transduced chlC::Tn10 to Ap^rTet^s on fusaric acidampicillin plates. Among these Ap^rTet^s potential chlC::Mu d1–8 fusions, a simple spot test identified the fusions that were closely linked to the Tn10 insertion in chlC. Out of 68 Ap^rTet^s isolates 7 chlC::Mu d1–8 fusions with a nitrate-induced Lac⁺ phenotype were obtained. When oxrA::Tn10 was transduced into these chlC::Mu d1–8 fusions, they became Lac^- even in the presence of nitrate, confirming that they were chlC::Mu d1–8 fusions.     

Enzyme kinetics of the prime K+ channel in the tonoplast of Chara: Selectivity and inhibition

Summary The prime potassium channel from the tonoplast of Chara corallina has been analyzed in terms of an enzyme kinetic model (Gradmann, Klieber & Hansen 1987, Biophys. J. 53:287) with respect to its selectivity for K⁺ over Rb⁺ and to its blockage by Cs⁺ and by Ca²⁺. The channel was investigated by patchclamp techniques over a range of membrane voltages (V _m , referred to an extracytoplasmic electrical potential of zero) from –200 mV to + 200 mV under various ionic conditions (0 to 300 mM K⁺, Rb⁺, Cs⁺, Ca²⁺, and Cl^–) on the two sides of isolated patches. The experimental data are apparent steady-state currentvoltage relationships under all experimental conditions used and amplitude histograms of the seemingly noisy open-channel currents in the presence of Cs⁺. The used model for K⁺ uniport comprises a reaction cycle of one binding site through four states, i.e., (1) K⁺-loaded and charged, facing the cytoplasm, (2) K⁺-loaded and charged facing the vacuole, (3) empty, facing the vacuole, and (4) empty, facing the cytoplasm. V _m enters the system in the form of a symmetric Eyring barrier between state 1 and 2. The numerical results for the individual rate constants are (in 10⁶s^–1 for zero voltage and 1 m substrate concentration): k ₁₂: 1,410, k ₂₁: 3,370, k ₂₃: 105,000, k ₃₂: 10,600, k ₃₄: 194, k ₄₃: 270, k ₄₁: 5,290, k ₁₄: 15,800. For the additional presence of an alternate transportee (here Rb⁺), the model can be extended in an analog way by another two states ((5) Rb⁺-loaded and charged, facing cytoplasm, and (6) Rb⁺-loaded and charged, facing vacuole) and six more rate constants (k ₄₅: 300, k ₅₄: 240, k ₅₆: 498, k ₆₅: 4,510, k ₆₃: 4,070, k ₃₆: 403). This six-state model with its unique set of fourteen parameters satisfies the complete set of experimental data. If the competing substrate can be bound but not translocated (here Cs⁺ and Ca²⁺), k ₅₆ and k ₆₅ of the model are zero, and the stability constants K _cyt (= k ₃₆/k ₆₃) and K _vac (= k ₄₅/k ₅₄) turn out to be K _cyt(Ca²⁺): 250 m ^–1 · exp(V _m /(64 mV)), k _vac(Ca²⁺): 10 m ^–1 · exp(–V _m /(66 mV)), K _cyt(Cs⁺): 0, and K _vac(Cs⁺): 46 m ^–2 · exp(–V _m /(12.25 mV)). With the assumption that the current fluctuations in the presence of Cs⁺ consist of incompletely resolved, short periods of complete openings and complete closures, the amplitude histograms of the noisy open channel currents can be described by a beta distribution, yielding the rate constants for binding (92 · 10⁶ sec^–1 · m ^–2 · exp(–V _m /(22.5 mV)) and debinding (2, 10⁶ sec^–1 · m ^–2 · exp(V _m /(22.5 mV)) of Cs⁺ to the vacuolar side of the channel as functions of the [Cs⁺] and of V _m . Considering these data and those from the literature, an asymmetry of the channel can be assessed, with a high charge density at the cytoplasmic side (Eisenman-series Nr. XI) and a low charge density at the vacuolar side (Eisenman-series Nr. I). Furthermore, the results provide an example for intimate linkage between conduction and switching of a channel.This work has been supported by the Deutsche Forschungsgemeinschaft.     

Mean-square hydrophobic moment for partially helical polypeptides

A mean-square helical hydrophobic moment, 〈h²〉, is defined for polypeptides in analogy to the mean-square dipole moment, 〈μ²〉, for polymer chains. For a freely jointed polymer chain, 〈μ²〉 is given by Σm, where m_i denotes the dipole moment associated with bond i. In the absence of any correlations in the hydrophobic moments of individual amino acid residues in the helix, 〈h²〉 is specified by ΣH, where H_i denotes the hydrophobicity of residue i. The tendency for correlations in orientations of residue hydrophobic moments in helices therefore dictates the size of 〈h²〉/〈H²〉, where 〈H²〉 denotes the average value of ΣH for all helices. The value of 〈h²〉/〈H²〉 will be greater than one in amphiphilic helices. A necessary prerequisite for this diagnostic usage of 〈h²〉/〈H²〉 is that the residue hydrophobic moment be oriented prependicular to the principal axis of the helix. Matrix-generation schemes are formulated that permit rapid evaluation of 〈h²〉 and 〈H²〉. The behavior of 〈h²〉/〈H²〉 is illustrated by calculations performed for model sequential copolypeptides.     

Optimization of growth conditions and fatty acid analysis for three freshwater diatom isolates

Diatoms are a group of highly abundant and diverse aquatic algae species. They contain high lipid content along with many bioactive compounds that can be exploited for biotechnological applications. Despite these attractive attributes, diatoms are underrepresented in production projects due to difficulties in their cultivation. To optimize the growth of three freshwater diatom isolates, Cyclotella sp., Synedra sp. and Navicula sp., an orthogonal assay on N, P, Si and Fe, as well as temperature and pH, was designed using traditional single‐factor tests. We also studied the effect of using nanosilica as an alternate Si source on growth and found that the diatom isolates studied achieved their highest growth rates under different combinations of nutrient and environmental conditions. Silica had the greatest influence on growth, followed by phosphate and iron. The optimized growth conditions for Synedra sp. were N: 30 mg L^?1, P: 3 mg L^?1, Si: 14.8 mg L^?1, Fe: 0.448 mg L^?1, temperature 25°C and pH 8. For Navicula sp.: N: 20 mg L^?1, P: 2.5 mg L^?1, Si: 19.7 mg L^?1, Fe: 0.112 mg L^?1, temperature 30°C and pH 7.5–8. For Cyclotella sp.: N: 20 mg L^?1, P: 2.5 mg L^?1, Si: 19.7 mg L^?1, Fe: 0.448 mg L^?1, temperature 30°C and pH 7.5–8. Nano silica negatively affected growth in Navicula sp. and Cyclotella sp., but no such effect was observed in Synedra sp. Fatty acid profiling showed C16:0, C16:1(n ? 7), C18:0 and C20:5(n ? 3) as major fatty acids, with no significant differences in fatty acid methyl ester profiles between traditional and modified media. This work gives us a new insight into the growth requirements of freshwater diatom species, which are less studied than marine species.     

Effect of ogt expression on mutation induction by methyl-, ethyl- and propylmethanesulphonate in Escherichia coli K12 strains

We have previously reported the isolation of an Escherichia coli K12 mutant that is extremely sensitive to mutagenesis by low doses of ethylating agents. We now show by Southern analysis that the mutation involves a gross deletion covering at least the ogt and fnr genes and that no O⁶-alkylguanine-DNA-alkyltransferase activity is present in cell-free extracts of an ada::Tn10 derivative of these bacteria. Confirmation that sensitisation to ethylation-induced mutagenesis was attributable to ogt and not to any other loci covered by the deletion was obtained by constructing derivatives. Thus an ogt::kan^r disruption mutation was introduced into the parental ogt ⁺ bacteria, and the ogt::kan^r mutation was then eliminated by cotransduction of ogt ⁺ with the closely linked Tet ^r marker (zcj::Tn10). The (ogt-fnr) deletion or ogt::kan^r disruption mutants were highly sensitive to ethyl methanesulphonate-induced mutagenesis, as measured by the induction of forward mutations to l-arabinose resistance (Ara¹). Furthermore, the number of Ara^r mutants increased linearly with dose, unlike the case in ogt ⁺ bacteria, which had a threshold dose below which no mutants accumulated. Differences in mutability were even greater with propyl methanesulphonate. Overproduction of the ogt alkyltransferase from a multicopy plasmid reduced ethylmethanesulphonate-induced mutagenesis in the ogt mutant strains and also methylmethanesulphonate mutagenesis in ada ^– bacteria. A sample of AB 1157 obtained from the E. coli K12 genetic stock centre also had a deletion covering the ogt and fnr genes. Since such deletions greatly influence the mutagenic responses to alkylating agents, a survey of the presence of the ogt gene in the E. coli K12 strain being used is advisable.     

Analysis of the frequency of inheritance of transposed Ds elements in Arabidopsis after activation by a CaMV 35S promoter fusion to the Ac transposase gene

The Ac/Ds transposon system of maize shows low activity in Arabidopsis. However, fusion of the CaMV 35S promoter to the transposase gene (35S::TPase) increases the abundance of the single Ac mRNA encoded by Ac and increases the frequency of Ds excision. In the experiments reported here it is examined whether this high excision frequency is associated with efficient re-insertion of the transposon. This was measured by using a Ds that carried a hygromycin resistance gene (HPT) and was inserted within a streptomycin resistance gene (SPT). Excision of Ds therefore gives rise to streptomycin resistance, while hygromycin resistance is associated with the presence of a transposed Ds or with retention of the element at its original location. Self-fertilisation of most individuals heterozygous for Ds and 35S::TPase produced many streptomycin-resistant (strep^r) progeny, but in many of these families a small proportion of strep^r seedlings were also resistant to hygromycin (hyg^r). Nevertheless, 70% of families tested did give rise to at least one strep^r, hyg^r seedling, and over 90% of these individuals carried a transposed Ds. In contrast, the Ac promoter fusion to the transposase gene (Ac::TPase) produced fewer strep^rhyg^r progeny, and only 53% of these carried a transposed Ds. However, a higher proportion of the strep^r seedlings were also hyg^r than after activation by 35S::TPase. We also examined the genotype of strep^r, hyg^r seedlings and demonstrated that after activation by 35S::TPase many of these were homozygous for the transposed Ds, while this did not occur after activation by Ac::TPase. From these and other data we conclude that excisions driven by 35S::TPase usually occur prior to floral development, and that although a low proportion of strep^r progeny plants inherit a transposed Ds, those that do can be efficiently selected with an antibiotic resistance gene contained within the element. Our data have important implications for transposon tagging strategies in transgenic plants and these are discussed.     

Molecular dynamics simulations of certain mutant peptide models from staphylococcal nuclease reveal that initial hydrophobic collapse associated with turn propensity drives β‐hairpin folding

An important nucleation event during the folding of staphylococcal nuclease involves the formation of a β‐hairpin by the sequence ²¹DTVKLMYKGQPMTFR³⁵. Earlier studies show that the turn sequence ‘YKGQP’ has an important role in the folding of this β‐hairpin. To understand the active or passive nature of the turn sequence ‘YKGQP’ in the folding of the aforementioned β‐hairpin sequence, we studied glycine mutant peptides Ac‐²DTVKLMYGGQPMTFR¹⁶‐NMe (K9G:15), Ac‐²DTVKLMYKGGPMTFR¹⁶‐NMe (Q11G:15), Ac‐²DTVKLMYGGGPMTFR¹⁶‐NMe (K9G/Q11G:15), and Ac‐²DTVKLMGGGGGMTFR¹⁶‐NMe (penta‐G:15) by using molecular dynamics simulations, starting with two different unfolded states, polyproline II and extended conformational forms. Further, 5mer mutant turn peptides Ac‐²YGGQP⁶‐NMe (K3G:5), Ac‐²YKGGP⁶‐NMe (Q5G:5), Ac‐²YGGGP⁶‐NMe (K3G/Q5G:5), and Ac‐²GGGGG⁶‐NMe (penta‐G:5) were also studied individually. Our results show that an initial hydrophobic collapse and loop closure occurs in all 15mer mutants, but only K9G:15 mutant forms a stable native‐like β‐hairpin. In the other 15mer mutants, the hydrophobic collapsed state would not proceed to β‐hairpin formation. Of the different simulations performed for the penta‐G:15 mutant, in only one simulation a nonnative β‐hairpin conformation is sampled with highly flexible loop region (⁸GGGGG¹²), which has no specific conformational preference as a 5mer. While the sequence ‘YGGQP’ in the K3G:5 simulation shows relatively higher β‐turn propensity, the presence of this sequence in K9G:15 peptide seems to be driving the β‐hairpin formation. Thus, these results seem to suggest that for the formation of a stable β‐hairpin, the initial hydrophobic collapse is to be assisted by a turn propensity. Initial hydrophobic collapse alone is not sufficient to guide β‐hairpin formation. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Phenotypic differences between Drosophila Alzheimer’s disease models expressing human Aβ42 in the developing eye and brain

Drosophila melanogaster expressing amyloid-β42 (Aβ42) transgenes have been used as models to study Alzheimer’s disease. Various Aβ42 transgenes with different structures induce different phenotypes, which make it difficult to compare data among studies which use different transgenic lines. In this study, we compared the phenotypes of four frequently used Aβ42 transgenic lines, UAS-Aβ42^2X, UAS-Aβ42^BL33770, UAS-Aβ42^11C39, and UAS-Aβ42^H29.3. Among the four transgenic lines, only UAS-Aβ42^2X has two copies of the upstream activation sequence-amyloid-β42 (UAS-Aβ42) transgene, while remaining three have one copy. UAS-Aβ42^BL33770 has the 3′ untranslated region of Drosophila α-tubulin, while the others have that of SV40. UAS-Aβ42^11C39 and UAS-Aβ42^H29.3 have the rat pre-proenkephalin signal peptide, while UAS-Aβ42^2X and UAS-Aβ42^BL33770 have that of the fly argos protein. When the transgenes were expressed ectopically in the developing eyes of the flies, UAS-Aβ42^2X transgene resulted in a strongly reduced and rough eye phenotype, while UAS-Aβ42^BL33770 only showed a strong rough eye phenotype; UAS-Aβ42^H29.3 and UAS-Aβ42^11C39 had mild rough eyes. The levels of cell death and reactive oxygen species (ROS) in the eye imaginal discs were consistently the highest in UAS-Aβ42^2X, followed by UAS-Aβ42^BL33770, UAS-Aβ42^11C39, and UAS-Aβ42^H29.3. Surprisingly, the reduction in survival during the development of these lines did not correlate with cell death or ROS levels. The flies which expressed UAS-Aβ42^11C39 or UAS-Aβ42^H29.3 experienced greatly reduced survival rates, although low levels of ROS or cell death were detected. Collectively, our results demonstrated that different Drosophila AD models show different phenotypic severity, and suggested that different transgenes may have different modes of cytotoxicity.

Abbreviations: Aβ42: amyloid-β42; AD: Alzheimer’s disease; UAS: upstream activation sequence     

Genetic complementation of rhizobial nod mutants with Frankia DNA: artifact or reality?

Two divergent reports have been published on the genetic complementation of rhizobial nod mutants using Frankia DNA. In 1991 putative Frankia cosmid library clones were reported to restore normal nodulation properties to Rhizobium leguminosarum biovar viciaenodD::Tn5, but no supporting sequence data were published. In 1992 a second group reported a failure to find any evidence of functional complementation of various rhizobial nod mutants by Frankia DNA (nodA, nodB and nodC). Complementation tests of nine Nod⁻ R. leguminosarum bv. viciae or Sinorhizobium meliloti Tn5 mutants (nodA ⁻ , nodB ⁻ , nodC ⁻ , nodD ⁻ , nodF  ⁻ , nodL ⁻ , nodH ⁻ ) were thus performed using a Frankia gene library in pLAFR3 to clarify this situation. Rhizobial transconjugants obtained by tri-parental matings were screened for restoration of the nodulation phenotype on their host plants, Vicia sativa subsp. nigra or Medicago sativa. Nodulation was observed on plants inoculated with transconjugants of the R. leguminosarum bv. viciaenodC::Tn5 mutant. The Nod⁺ rhizobial transconjugants were isolated and analysed. The Nod⁺ phenotype of these transconjugants was found to be due to Tn5 excision/transposition. No functional complementation was found with any of the mutants used, suggesting that rhizobial complementation of nod mutants with Frankia DNA is unlikely to occur. Received: 17 April 1998 / Accepted: 22 July 1998     

Gene-directed mutagenesis on the chromosome of Bacillus subtilis 168

Summary We have devised a method whereby any mutagenized cloned DNA from Bacillus subtilis can be reinserted at the original site on the B. subtilis chromosome. The procedure depends on the accuracy and high frequency of homologous recombination between the B. subtilis chromosome and the DNA taken up by the cell. The method makes use of two drug resistance selection markers (the chloramphenicol resistance gene and the neomycin resistance gene) and a marker gene which functions as a catalyst. The utility of the method has been demonstrated using leuB and pro of B. subtilis as target gene and catalyst, respectively, and mutations such as leuB: : cat, leuB ^–, and pro: : neo constructed in vitro on the cloned DNA fragments. Transformation in sequential steps as (leuB ⁺ pro⁺)(leuB: : cat pro ⁺) (leuB ^– pro: : neo)(leuB ^– pro ⁺) resulted in a leuB ^– single mutant without affecting other regions of the B. subtilis chromosome (gene-directed mutagenesis). We also demonstrate that other single mutations such as metD ^– and pro ^–, as well as the double mutation leuB ^– pro ^– can be introduced by the same procedure. In principle, true isogenies with multiple mutations can be constructed by the method described in this paper. Furthermore, the procedure should be generally applicable to any organisms in which homologous recombination is proficient.     

Isolation of a formamidopyrimidine-DNA glycosylase (fpg) mutant of Escherichia coli K12

Summary The fpg ⁺ gene of Escherichia coli coding for formamidopyrimidine-DNA glycosylase was previously cloned on a multicopy plasmid. The plasmid copy of the fpg ⁺ gene was inactivated by cloning a kanamycin resistance gene into the open reading frame, yielding the fpg-1:: Kn^r mutation. This mutation was transferred to the chromosome in the following steps: (i) linearization of the plasmid bearing the fpg-1::Kn^r mutation and transformation of competent bacteria (recB recC sbcB); (ii) selection for chromosomal integration of the fpg-1::Kn^r mutation; (iii) phage P1 mediated transduction of the fpg-1::Kn^r mutation in the AB1157 background. The resulting fpg ^- mutant exhibited no detectable Fapy-DNA glycosylase activity in crude lysates. The insertion mutation was localized by means of genetic crosses between mtl and pyrE, at 81.7 min on the E. coli linkage map. Sequence analysis confirmed this mapping and further showed that fpg is adjacent to rpmBG in the order fpg, rpmGB, pyrE. The formamidopyrimidine-DNA glycosylase defective strain does not show unusual sensitivity to the following DNA damaging treatments: (i) methylmethanesulfonate, (ii) N-methyl-N-nitro-N-nitrosoguanidine, (iii) ultraviolet light, (iv) -radiation. The fpg gene is neither part of the SOS regulon nor the adaptive response to alkylating agents.     

Production studies on protozoa

Summary In the river Saale and in the terrestrial moss Mnium cuspidatum Leyss. in 1974/75 the annual production of Testacea and loricate ciliated protozoa were investigated.The production was estimated in the Saale-Aufwuchs on a -meso ... oligosaprobic (Kaulsdorf, Thuringia, GDR) and on a -mesosaprobic (Rothenstein, Thuringia, GDR) area of the river. The mosses were investigated in a forest near Jena.The production was estimated on slides and in special productionchambers; the time of exposure was 2 weeks. Investigations concerned annual production of individuals and biomass, the ratio of annual production/standing crop (P/B), numbers of generations per year (G) and mortality (M%/d). In the mosses, the rainfall modified the production and dislocation of the protozoa.The values for production are: Aufwuchs Saale (-meso... oligosaprobic): 24·10⁶ i/m²·a (=1,0 g/m²·a=79·10³ i/m²·d); P/B: 12.6. Aufwuchs Saale (-mesosaprobic): 3.2·10⁶ i/m²·a (=0.35 g/m²·a=81·10³ i/m²·d); P/B: 34.9; G: 22; M: 5%/d. Moss: 145·10⁶ i/m²·a (=0.11 g/m²·a=40.6·10³ i/m²·d); P/B: 8.1; G: 16.5; M: 3.0%/d.     

Effect of Voluntary Exercise on Genetically Obese Cpefat/fat Mice: Quantitative Proteomics of Serum

Objective: To compare the effect of voluntary exercise on body weight, food consumption, and levels of serum proteins between wild‐type and carboxypeptidase E‐deficient (Cpe^fat/fat) mice. Research Methods and Procedures: Study 1 consisted of three groups of female mice: Cpe^fat/fat mice with continuous access to exercise wheels for 3 weeks (n = 4); wild‐type C57BKS mice with access to exercise wheels for 3 weeks (n = 4); and sedentary Cpe^fat/fat mice (n = 3). Activity, body weight, and food consumption were monitored for this period and a subsequent 9‐week period without exercise wheels. Study 2 consisted of four groups of male mice (n = 6 to 7 each): Cpe^fat/fat mice with exercise wheels, wild‐type mice with exercise wheels, and Cpe^fat/fat and wild‐type mice without exercise wheels. Body weight and food consumption were measured over 4 weeks. Sera were collected, and the protein profile was determined by 2‐dimensional gel electrophoresis and mass spectrometry. Results: Cpe^fat/fat mice were moderately hyperphagic but lost weight during the initial exercise period because of greater energy expenditure. The effect of exercise was temporary, and the mice gained weight after the second week. Several serum proteins were found to be altered by exercise: haptoglobin was decreased by exercise in Cpe^fat/fat mice, and several kallikreins were increased by exercise in wild‐type mice. Discussion: The access to exercise wheels provided an initial weight loss in Cpe^fat/fat mice, but this effect was offset by elevated food consumption. The serum proteomics results indicated that Cpe^fat/fat and wild‐type mice differed in their response to exercise.     

Diversity of the Carbohydrate Composition of the Antigenic Polysaccharides of Proteobacteria of the Genera Pseudoalteromonasand Marinomonas

The sugar analysis of the glycans of the type strains of marine proteobacteria of the genera Pseudoalteromonasand Marinomonas—Pseudoalteromonas atlanticaIAM12927^T, P. aurantiaNCIMB 2033^T, P. citreaATCC 29719^T, P. elyakoviiKMM 162^T, P. espejianaATCC 29659^T, P. piscicidaNCIMB 645^T, P. tetraodonisIAM 14160^T, Marinomonas communisATCC 27118^T, and M. vagaATCC 27119^T—showed that they contain glucose, galactose, galactosamine, glucosamine, fucose, rhamnose, mannose, heptose, 2-keto-3-deoxyoctonate (KDO), uronic acids, colitose (3,6-dideoxy-L-xylo-hexose), and 6-deoxy-L-talose. The carbohydrate composition of the antigenic polysaccharides (PSs) of P. elyakoviiKMM 162^Tand P. espejianaATCC 29659^Tdepended on the type and the concentration of carbohydrate substrates in the nutrient media. The molar proportion between rhamnose, glucose, and galactose (ca. 1 : 0.3 : 2) in the PS of P. elyakoviiKMM 162^Twas almost the same in the media lacking carbohydrates or containing glucose or galactose at a concentration of 1 g/l. At the same time, the molar proportion between fucose, glucose, galactose, galactosamine, and glucosamine (ca. 1 : 1 : 1 : 2 : 0.5) in the PS of P. espejianaATCC 29659^Tdepended on the presence and the concentration of carbohydrate substrates in the medium. A high concentration of glucose in the medium (30 g/l) brought about a rise in the content of glucose in PSs (9-fold for the PS of P. elyakoviiKMM 162^Tand 4.6-fold for the PS of P. espejianaATCC 29659^T) and led to a decrease in the content of other carbohydrates. The cultivation of these two strains at a lactose concentration of 30 g/l resulted in their PSs containing glucose and galactose in about equal proportions (ca. 1 : 1 in the case of P. espejianaATCC 29659^Tand ca. 2.1 : 1.7 in the case of P. elyakoviiKMM 162^T).     

Temporal variation in body composition (C : N) helps explain seasonal patterns of zooplankton δ13C

1. The stable carbon isotope ratio δ¹³C is a useful tracer of energy flow in lake food webs, and the zooplankton signature is commonly used to establish a baseline for the pelagic habitat. However, sources of temporal variability in the δ¹³C of different zooplankton taxa are rarely considered. 2. Here, we investigate to what extent temporal variation in the δ¹³C of particulate organic matter (POM) (<41 μm) and the C : N of zooplankton can explain the temporal variability in δ¹³C of freshwater zooplankton. We compare temporal patterns of δ¹³C and C : N for Daphnia, Hesperodiaptomus franciscanus and Leptodiaptomus tyrelli over a 6‐month period at four sites in two oligotrophic lakes. 3. In all three taxa, seasonal variation in zooplankton C : N explained more of the variation in zooplankton δ¹³C than did the δ¹³C of POM. This suggests that variation in the lipid content of zooplankton can strongly influence temporal variation of δ¹³C in zooplankton. 4. Using these data, we evaluate procedures that estimate the δ¹³C of only the non‐lipid component of zooplankton. If zooplankton lipids are primarily dietary in origin, than extracting lipids or ‘normalising’δ¹³C based on C : N will exclude a major dietary source, and therefore may be inappropriate. 5. We conclude that temporal variation in body composition (C : N) of zooplankton can significantly influence the temporal variation of zooplankton δ¹³C signatures.     

Functional defect of peripheral neutrophils in mice with induced deletion of CXCR2

Type 2 CXC chemokine receptor CXCR2 plays roles in development, tumorigenesis, and inflammation. CXCR2 also promotes demyelination and decreases remyelination by actions toward hematopoietic cells and nonhematopoietic cells. Germline CXCR2 deficient (Cxcr2^‐/‐) mice reported in 1994 revealed the complexity of CXCR2 function and its differential expression in varied cell‐types. Here, we describe Cxcr2^fl/fl mice for which the targeting construct was generated by recombineering based on homologous recombination in E. coli. Without recombination Cxcr2^fl/fl mice have CXCR2 expression on neutrophils in peripheral blood, bone marrow and spleen. Cxcr2^fl/fl mice were crossed to Mx‐Cre mice in which Cre recombinase is induced by Type I interferons, elicited by injection with polyinosinic‐polycytidylic acid (poly(I:C)). CXCR2‐deficient neutrophils were observed in poly(I:C) treated Cxcr2^fl/fl::Mx‐Cre⁺ (Cxcr2‐CKO) mice, but not in poly(I:C) treated Cxcr2^f//+::Mx‐Cre⁺ mice. CXCR2 deletion was mainly observed peripherally but not in the CNS. Cxcr2‐CKO mice showed impaired neutrophil migration in sterile peritonitis. Cxcr2‐CKO mice reported here will provide a genetic reagent to dissect roles of CXCR2 in the neutrophil granulocyte lineage. Furthermore Cxcr2^fl/fl mice will provide useful genetic models to evaluate CXCR2 function in varied cell populations. genesis 51:587–595. © 2013 Wiley Periodicals, Inc.     

Esterase-2 in Ephestia kühniella. II. Tissue-specific patterns

Esterase-2 polymorphism has been investigated quantitatively. Staining intensities of the homodimer bands mm, ff, and ss are not equally expressed but found in relative activities of 1:0.5:0.7 in fat bodies and 1:1:3 in testes and gut walls. Relative intensities of the parental bands are binomially distributed in the three-banded patterns of heterozygotes in an exactly tissue-specific manner. Organ-specific proportions of relative activity remain constant throughout postembryonic development. Among reasons which may influence genesis of allele- and organ-specific activities of homodimer bands in homozygotes and may lead to asymmetrical distribution of intensities in the patterns of heterozygotes, a hypothesis of differential allelic activity is discussed, according to which the structural alleles 2 ^m , 2 ^f , and 2 ^s are closely linked to the alleles RG ^a , RG ^b , and RG ^c of a controlling gene.This work was supported by the Deutsche Forschungsgemeinschaft and submitted with Part I in partial fulfillment of the requirements for the degree of Dr.-habil. at the University of Tübingen.     

Competition between effective and ineffective bradyrhizobia nodulating peanuts (Arachis hypogaea)1

The competitive ability of effective and ineffective isolates of Bradyrhizobium to form nodules on four peanut (Arachis hypogaea) genotypes from which they were originally isolated was investigated in a greenhouse study. Pregerminated seeds of each genotype were inoculated with five ratios of ineffective effective bradyrhizobia: 10⁴:0, 10⁴:10² 10⁴:10⁴ 10²:10⁴ and 0:10⁴. Plants were harvested 35 days after planting and nitrogenase activity, total nodule number per plant, plant shoot dry weight and the proportion of nodules formed by each Bradyrhizobium isolate were determined. Nodulation suppression occurred on plants inoculated with the mixed cultures, especially on those inoculated with the 10⁴:10² ineffective effective inoculum ratio. More than 85% of the nodules on plants inoculated with the 10⁴:10⁴ and 10²:10⁴ ineffective effective Bradyrhizobium mixtures were formed by the effective isolates. The number of effective nodules per plant, nitrogenase activity and plant shoot dry matter production all increased with the proportion of the effective bradyrhizobia in the inocula.