Bacterial-based systems for expression and purification of recombinant Lassa virus proteins of immunological relevance

<p>Abstract</p> <p>Background</p> <p>There is a significant requirement for the development and acquisition of reagents that will facilitate effective diagnosis, treatment, and prevention of Lassa fever. In this regard, recombinant Lassa virus (LASV) proteins may serve as valuable tools in diverse antiviral applications. Bacterial-based systems were engineered for expression and purification of recombinant LASV nucleoprotein (NP), glycoprotein 1 (GP1), and glycoprotein 2 (GP2).</p> <p>Results</p> <p>Full-length NP and the ectodomains of GP1 and GP2 were generated as maltose-binding protein (MBP) fusions in the Rosetta strains of <it>Escherichia coli </it>(<it>E. coli</it>) using pMAL-c2x vectors. Average fusion protein yields per liter of culture for MBP-NP, MBP-GP1, and MBP-GP2 were 10 mg, 9 mg, and 9 mg, respectively. Each protein was captured from cell lysates using amylose resin, cleaved with Factor Xa, and purified using size-exclusion chromatography (SEC). Fermentation cultures resulted in average yields per liter of 1.6 mg, 1.5 mg, and 0.7 mg of purified NP, GP1 and GP2, respectively. LASV-specific antibodies in human convalescent sera specifically detected each of the purified recombinant LASV proteins, highlighting their utility in diagnostic applications. In addition, mouse hyperimmune ascitic fluids (MHAF) against a panel of Old and New World arenaviruses demonstrated selective cross reactivity with LASV proteins in Western blot and enzyme-linked immunosorbent assay (ELISA).</p> <p>Conclusion</p> <p>These results demonstrate the potential for developing broadly reactive immunological assays that employ all three arenaviral proteins individually and in combination.</p

Genetic Variation and Breeding Potential of Phytate and Inorganic Phosphorus in a Maize Population

Seed P is predominantly bound in the organic compound phytate, which makes the bioavailability of P low for monogastric animals fed maize (Zea mays L.)-based diets. Decreasing phytate and increasing inorganic P (Pi, an available form of P) concentrations in maize grain would be desirable to help ameliorate environmental problems associated with high P in feces. Our objective was to investigate the potential of improving the P profile of maize grain through breeding and selection. Ninety S1 families from the BS31 population were evaluated at two locations for phytate, Pi, and other grain quality and agronomic traits. Phytate concentrations ranged from 1.98 to 2.46 g kg−1, and the broad-sense heritability (H) was relatively low (0.60). Both genetic variance and H (0.84) were much greater for Pi Few unfavorable genetic correlations were observed between either Pi or phytate and other key economic traits. Also, selection differentials of multiple trait indices indicated that the P profile of maize grain and grain yield and moisture could be improved simultaneously. Many cycles of selection will be needed, however, to reach desirable phytate and Pi concentrations, especially when selecting for multiple traits. Regardless, our results are encouraging given that the families evaluated were related S1 families and the number of families was relatively small

Genetic mapping of quantitative trait loci for end-use quality and grain minerals in hard red winter wheat

To meet the demands of different wheat-based food products, traits related to end-use quality become indispensable components in wheat improvement. Thus, markers associated with these traits are valuable for the timely evaluation of protein content, kernel physical characteristics, and rheological properties. Hereunder, we report the mapping results of quantitative trait loci (QTLs) linked to end-use quality traits. We used a dense genetic map with 5199 SNPs from a 90K array based on a recombinant inbred line (RIL) population derived from ‘CO960293-2’/‘TAM 111’. The population was evaluated for flour protein concentration, kernel characteristics, dough rheological properties, and grain mineral concentrations. An inclusive composite interval mapping model for individual and across-environment QTL analyses revealed 22 consistent QTLs identified in two or more environments. Chromosomes 1A, 1B, and 1D had clustered QTLs associated with rheological parameters. Glu-D1 loci from CO960293-2 and either low-molecular-weight glutenin subunits or gliadin loci on 1A, 1B, and 1D influenced dough mixing properties substantially, with up to 34.2% of the total phenotypic variation explained (PVE). A total of five QTLs associated with grain Cd, Co, and Mo concentrations were identified on 3B, 5A, and 7B, explaining up to 11.6% of PVE. The results provide important genetic resources towards understanding the genetic bases of end-use quality traits. Information about the novel and consistent QTLs provided solid foundations for further characterization and marker designing to assist selections for end-use quality improvements.Horticulture and Landscape Architectur

Nutrient partitioning and grain yield of \u3ci\u3eTaNAM\u3c/i\u3e-RNAi wheat under abiotic stress

Aims — Decreased expression of TaNAM genes by RNAi results in delayed senescence and decreased grain protein, iron, and zinc concentrations. Here, we determined whether NAM expression level alters onset of senescence under stress conditions, whether delayed senescence in the TaNAM-RNAi line resulted in improved tolerance to post-anthesis abiotic stress, and determined the effects of post-anthesis abiotic stress on N and mineral remobilization and partitioning to grain. Methods — Greenhouse-grown WT and TaNAM-RNAi wheat were characterized in two studies: three levels of N fertility or water limitation during grain fill. Studies were conducted under both optimal and heat stress temperatures. Senescence onset was determined by monitoring flag leaf chlorophyll. Results — Under optimal temperatures, TaNAM-RNAi plants had a yield advantage at lower N. TaNAM-RNAi plants had delayed senescence relative to the WT and lower grain protein and mineral concentrations, N remobilization efficiency, and partitioning of N and most minerals to grain. Conclusions — Nutritional quality of TaNAM-RNAi grain was consistently lower than WT. Delayed senescence of TaNAM-RNAi plants provided a yield advantage under optimal temperatures but not under water or heat stress. Discovery of specific NAM protein targets may allow separation of the delayed senescence and nutrient partitioning traits, which could be used for improvement of wheat. Includes supplementary materials

Registration of ‘Juniper’ Wheat

‘Juniper’ (Reg. No. CV-1021, PI 639951) is a hard red winter wheat (Triticum aestivum L.) developed by the Idaho Agricultural Experiment Station and released in February 2006. Juniper was tested under the experimental numbers A91013W-1 and IDO575. It is a full-stature wheat released for very low rainfall, crop-fallow rotations of the Intermountain West and was released for its improved resistance to stripe rust (Puccinia striiformis Westend) and dwarf bunt (Tilletia controversa Kühn in Rabenh) and bread-baking quality compared with current full-stature cultivars. Juniper had an average yield in rainfed trials of 3290 kg ha−1, compared with the other tall cultivars for this region, ‘Weston’ and ‘Bonneville’, which had grain yields of 3050 and 3180 kg ha−1, respectively. Weston has undesirable baking quality due to short dough mixing time. Juniper has nearly a full minute longer mixograph mixing time compared with Weston (p \u3c 0.01). Juniper also has similar snow mold tolerance (causal organism Typhula spp.) to Bonneville without the undesirable characteristic of late heading date and maturity. Juniper heads 3 d earlier than Bonneville