Absence of a 23 kd protein in testes of testicular feminization rat

Cryptorchid testes of testicular feminization rats are very low in zinc in spite of normal zinc status of the animals. Analysis of the cytosol of the cryptorchid testes by gel permeation chromatography showed decreased zinc binding by proteins eluted at fractions corresponding to 30,000 dalton. Further analysis by sodium dodecylsulphate polyacrylamide gel electrophoresis indicated the absence of a protein with molecular weight of 23,000.     

Changes in Bacterial and Fungal Communities across Compost Recipes,Preparation Methods,and Composting Times

Compost production is a critical component of organic waste handling, and compost applications to soil are increasingly important to crop production. However, we know surprisingly little about the microbial communities involved in the composting process and the factors shaping compost microbial dynamics. Here, we used high-throughput sequencing approaches to assess the diversity and composition of both bacterial and fungal communities in compost produced at a commercial-scale. Bacterial and fungal communities responded to both compost recipe and composting method. Specifically, bacterial communities in manure and hay recipes contained greater relative abundances of Firmicutes than hardwood recipes with hay recipes containing relatively more Actinobacteria and Gemmatimonadetes. In contrast, hardwood recipes contained a large relative abundance of Acidobacteria and Chloroflexi. Fungal communities of compost from a mixture of dairy manure and silage-based bedding were distinguished by a greater relative abundance of Pezizomycetes and Microascales. Hay recipes uniquely contained abundant Epicoccum, Thermomyces, Eurotium, Arthrobotrys, and Myriococcum. Hardwood recipes contained relatively abundant Sordariomycetes. Holding recipe constant, there were significantly different bacterial and fungal communities when the composting process was managed by windrow, aerated static pile, or vermicompost. Temporal dynamics of the composting process followed known patterns of degradative succession in herbivore manure. The initial community was dominated by Phycomycetes, followed by Ascomycota and finally Basidiomycota. Zygomycota were associated more with manure-silage and hay than hardwood composts. Most commercial composters focus on the thermophilic phase as an economic means to insure sanitation of compost from pathogens. However, the community succeeding the thermophilic phase begs further investigation to determine how the microbial dynamics observed here can be best managed to generate compost with the desired properties.     

Dissecting the telomere region of barley chromosome 5HL using rice genomic sequences as references: new markers for tracking a complex region in breeding

The terminal region of barley chromosome 5HL controls malt extract, diastatic power, free amino acid nitrogen, alpha-amylase activity, seed dormancy and pre-harvest sprouting. Comparative analysis of the barley and rice maps has established that the terminal region of barley chromosome 5HL is syntenic to rice chromosome 3L near the telomere end. The rice BAC (Bacterial Artificial Chromosome) sequences covering the region of chromosome 3L were used to search barley expressed sequenced tags database. Thirty-three genes were amplified by PCR (polymerase chain reaction) with the primers designed from barley ESTs (expressed sequence tag). Comparison of the sequences of the PCR generated DNA fragments revealed polymorphisms including single nucleotide polymorphism (SNP), insertions or deletions between the barley varieties. Seven new PCR based molecular markers were developed and mapped within 10 cM in three doubled haploid barley populations (Stirling × Harrington, Baudin × AC Metcalfe and Chebec × Harrington). The mapped genes maintain the micro-syntenic relationship between barley and rice. These gene specific markers provide simple and efficient tools for germplasm characterization and marker-assisted selection for barley malting quality, and ultimately lead to isolation and identification of the major gene(s) controlling multiple quality traits on barley chromosome 5HL.     

Discrimination of sunflower volatiles by the red sunflower seed weevil

Five principle monoterpenoid and other constituent volatile chemicals of sunflower heads were combined to resemble two lines of sunflower (Helianthus annuus L.): one U.S.D.A. standard line and one French line which was poorly visited by insects (Etievant et al., 1984). Field trials of attraction to red sunflower seed weevils (Smicronyx fulvus Le Conte, Coleoptera: Curculionidae) showed that one was clearly preferred over the other. The more attractive mixture contained -pinene, -pinene, limonene, camphene and bornyl acetate in a ratio resembling that of Flath et al. (1985) rather than that described by Etievant et al. (1984). One or two volatiles were deleted from the optimal blend but only mixtures of five volatiles showed the highest attraction. Substitution of sabinene, another volatile prominent in sunflower, for one of the five in the optimal blend also decreased attraction of seed weevils. When the monoterpenoid components and green leaf volatiles in the traps resembled the ratios of most of the prominent volatiles of sunflower, attraction was significantly greater than controls.     

Rapid determination of free proline for water-stress studies

Summary Proline, which increases proportionately faster than other amino acids in plants under water stress, has been suggested as an evaluating parameter for irrigation scheduling and for selecting drought-resistant varieties. The necessity to analyze numerous samples from multiple replications of field grown materials prompted the development of a simple, rapid colorimetric determination of proline. The method detected proline in the 0.1 to 36.0 moles/g range of fresh weight leaf material.     

Critical but distinct roles for the pleckstrin homology and cysteine-rich domains as positive modulators of Vav2 signaling and transformation

Vav2, like all Dbl family proteins, possesses tandem Dbl homology (DH) and pleckstrin homology (PH) domains and functions as a guanine nucleotide exchange factor for Rho family GTPases. Whereas the PH domain is a critical positive regulator of DH domain function for a majority of Dbl family proteins, the PH domains of the related Vav and Vav3 proteins are dispensable for DH domain activity. Instead, Vav proteins contain a cysteine-rich domain (CRD) critical for DH domain function. We evaluated the contribution of the PH domain and the CRD to Vav2 guanine nucleotide exchange, signaling, and transforming activity. Unexpectedly, we found that mutations of the PH domain impaired Vav2 signaling, transforming activity, and membrane association. However, these mutations do not influence exchange activity on Rac and only slightly affect exchange on RhoA and Cdc42. We also found that the CRD was critical for the exchange activity in vitro and contributed to Vav2 membrane localization. Finally, we found that phosphoinositol 3-kinase activation synergistically enhanced Vav2 transforming and signaling activity by stimulating exchange activity but not membrane association. In conclusion, the PH domain and CRD are mechanistically distinct, positive modulators of Vav2 DH domain function in vivo.     

Coencapsulation of irinotecan and floxuridine into low cholesterol-containing liposomes that coordinate drug release in vivo

A liposomal delivery system that coordinates the release of irinotecan and floxuridine in vivo has been developed. The encapsulation of floxuridine was achieved through passive entrapment while irinotecan was actively loaded using a novel copper gluconate/triethanolamine based procedure. Coordinating the release rates of both drugs was achieved by altering the cholesterol content of distearoylphosphatidylcholine (DSPC)/distearoylphosphatidylglycerol (DSPG) based formulations. The liposomal retention of floxuridine in plasma after intravenous injection was dramatically improved by decreasing the cholesterol content of the formulation below 20 mol%. In the case of irinotecan, the opposite trend was observed where increasing cholesterol content enhanced drug retention. Liposomes composed of DSPC/DSPG/Chol (7:2:1, mole ratio) containing co-encapsulated irinotecan and floxuridine at a 1:1 molar ratio exhibited matched leakage rates for the two agents so that the 1:1 ratio was maintained after intravenous administration to mice. The encapsulation of irinotecan was optimal when copper gluconate/triethanolamine (pH 7.4) was used as the intraliposomal buffer. The efficiency of irinotecan loading was approximately 80% with a starting drug to lipid molar ratio of 0.1/1. Leakage of floxuridine from the liposomes during irinotecan loading at 50 degrees C complicated the ability to readily achieve the target 1:1 irinotecan/floxuridine ratio inside the formulation. As a result, a procedure for the simultaneous encapsulation of irinotecan and floxuridine was developed. This co-encapsulation method has the advantage over sequential loading in that extrusion can be performed in the absence of chemotherapeutic agents and the drug/drug ratios in the final formulation can be more precisely controlled.     

pHG165: A pBR322 copy number derivative of pUC8 for cloning and expression

During the construction of the Messing pUC plasmid series, the rop(rom) gene of pBR322 which mediates the activity of RNAI was deleted. This has resulted in an elevated copy number for the pUC plasmids which makes the expression of beta-galactosidase activity constitutive in a host containing the Iqtss lac repressor. We describe the construction of a new series of vectors which retain the pUC multiple cloning site (MCS) but in which copy number control has been recovered. In addition, the lac alpha/lac promoter expression region has been inserted into a HpaI cassette. This facilitates the movement of recombinant DNA clones within the MCS. It also increases the complementation activity of the lac alpha peptide by an order of magnitude, allowing selection of recombinants by their Lac- phenotype on MacConkey agar.