Sequence and analysis of genomic sequences upstream of mei-3.

he Neurospora crassa mei-3 mutation causes sensitivity to various DNA damaging agents (Newmeyer and Galeazzi 1977 Genetics 85:461-487). We have recently cloned and mapped a genomic fragment capable of transforming mei-3 spheroplasts to wild type (Cheng et al. 1993 Mut. Res. 294:223-234). These experiments predicted the putative coding sequence of mei-3 or at least the region encompassing the mei-3 mutation. It was determined by homology that Mei-3 belongs to the RecA-like group of proteins (Bishop et al. 1992 Cell 69:439-456) which are intimately involved in the recombination process and had been previously identified only in prokaryotes. Since the identification of RecA-like proteins in Saccharomyces (Shinohara et al. 1992 Cell 69:457-470) and in Neurospora, our hypothesis that this important group of proteins may be highly conserved in other eukaryotes has been substantiated with the cloning of several Rec-A like proteins from mouse, chicken, lily, and human (Shinohara et al. 1993 Nat. Genet. 4:239-243). Using these data there is evidence that additional homology, upstream of our putative start site, to these other proteins exists (an additional 67% over 39 amino acids between Mei-3 and mouse Rad51: Figure 1). However, this region of homology lacks a start site or obvious splice sites. This might suggest the presence of an unidentified upstream start sequence and another exon without obvious splice sequences. In an attempt to address this possibility, we sequenced both strands of the previously unpublished genomic sequence, from -2519 to -271 bp upstream of the putative 5\u27 end of mei-3

The use of a nonradioactive probe in RFLP analysis of Neurospora crassa DNA

Our laboratory is investigating the use of nonradioactive alternatives for the synthesis of DNA probes used in hybridization experiments

Ligand binding and conformational dynamics of the E. coli nicotinamide nucleotide transhydrogenase revealed by hydrogen/deuterium exchange mass spectrometry

Nicotinamide nucleotide transhydrogenases are integral membrane proteins that utilizes the proton motive force to reduce NADP+ to NADPH while converting NADH to NAD+. Atomic structures of various transhydrogenases in different ligand-bound states have become available, and it is clear that the molecular mechanism involves major conformational changes. Here we utilized hydrogen/deuterium exchange mass spectrometry (HDX-MS) to map ligand binding sites and analyzed the structural dynamics of E. coli transhydrogenase. We found different allosteric effects on the protein depending on the bound ligand (NAD+, NADH, NADP+, NADPH). The binding of either NADP+ or NADPH to domain III had pronounced effects on the transmembrane helices comprising the proton-conducting channel in domain II. We also made use of cyclic ion mobility separation mass spectrometry (cyclic IMS-MS) to maximize coverage and sensitivity in the transmembrane domain, showing for the first time that this technique can be used for HDX-MS studies. Using cyclic IMS-MS, we increased sequence coverage from 68 % to 73 % in the transmembrane segments. Taken together, our results provide important new insights into the transhydrogenase reaction cycle and demonstrate the benefit of this new technique for HDX-MS to study ligand binding and conformational dynamics in membrane proteins

Students perceiving risk: a quantitative assessment on three South African university campuses

This paper documents risk as perceived by students in three South African universities—it focuses on risk encountered in everyday choices such as sexual behaviour, alcohol use and freedom of association.The project originated in qualitative research conducted over two years (2008- 2009) among Rhodes University (RU) Sociology students. A second—quantitative—phase began in 2010 when findings from the initial phase were used to guide students in designing and piloting a questionnaire. Pilot results then informed a final research iteration in which a second questionnaire was designed and administered at three universities.The final questionnaire proved robust and performed very well. Among the 1694 valid cases included in analyses, strong associations were found between the universities, respondents’ sex and their worldview. Exploratory factor analysis revealed four latent constructs underlying all responses: The existential experience of risk; Stress and pressure resulting from exposure to financial and emotional risk; Risks attached to the everyday university culture; and Promiscuity. These constructs explained 59.8% of the total variance in all observed cases (all respondents in all universities).Students’ perception of risk in their lives is complex and multi-dimensional. The research found considerable evidence for Beck’s claim that in late modernity there is increasing individualisation in people’s perception of risk. Also, the degrees to which specific risks resonate in student consciousness demonstrate that risk is—as Zinn claims—both constructed and real.Key Words: risk, student behaviour, existential experience, sexual violence, South AfricaRésuméCet article documente le risque perçu par les étudiants dans trois universités sud- africaines concentre-il un risque rencontré dans les choix de tous les jours tels que le comportement sexuel, la consommation d’alcool et la liberté de projet association. The origine dans la recherche qualitative Menée sur deux ans (2008-2009) entre Les étudiants de l’Université de Rhodes (RU) sociologie. Une deuxième phase quantitative a commencé en 2010 lorsque les résultats de la première phase ont été utilisés pour guider les élèves dans la conception et le pilotage d’un questionnaire. Résultats de pilote informe alors une itération de recherche final dans lequel un second questionnaire a été conçu et administré à trois universities.The questionnaire final s’est avéré robuste et très bien performé. Parmi les 1 694 cas valides incluses dans les analyses, de fortes associations ont été trouvées entre les universités, le sexe des répondants et de leur vision du monde. L’analyse factorielle exploratoire a révélé quatre construits latents sous-jacents toutes les réponses: L’expérience existentielle de risque; Le stress et la pression résultant de l’exposition au risque financier et émotionnel; Risques liés à la culture universitaire de tous les jours; et la promiscuité. Ces constructions ont expliqué 59,8% de la variance totale dans tous les-cas observés (tous les répondants dans toutes les universités). La perception du risque dans leur vie des étudiants est complexe et multidimensionnelle. La recherche a trouvé des preuves considérables pour la demande de Beck C’est à la fin de la modernité, il ya augmentation de l’individualisation dans la perception des gens de risque. En outre, les degrés dans laquelle les risques spécifiques résonnent dans la conscience des étudiants démontrent C’est le risque que les créances Zinn appliquées Bothan- construit et réel.Mots clés: risque, le comportement des élèves, l’expérience existentielle, la violence sexuelle, l’Afrique du Su

Bee-hawking by the wasp, Vespa velutina, on the honeybees Apis cerana and A. mellifera

The vespine wasps, Vespa velutina, specialise in hawking honeybee foragers returning to their nests. We studied their behaviour in China using native Apis cerana and introduced A. mellifera colonies. When the wasps are hawking, A. cerana recruits threefold more guard bees to stave off predation than A. mellifera. The former also utilises wing shimmering as a visual pattern disruption mechanism, which is not shown by A. mellifera. A. cerana foragers halve the time of normal flight needed to dart into the nest entrance, while A. mellifera actually slows down in sashaying flight manoeuvres. V. velutina preferentially hawks A. mellifera foragers when both A. mellifera and A. cerana occur in the same apiary. The pace of wasp-hawking was highest in mid-summer but the frequency of hawking wasps was three times higher at A. mellifera colonies than at the A. cerana colonies. The wasps were taking A. mellifera foragers at a frequency eightfold greater than A. cerana foragers. The final hawking success rates of the wasps were about three times higher for A. mellifera foragers than for A. cerana. The relative success of native A. cerana over European A. mellifera in thwarting predation by the wasp V. velutina is interpreted as the result of co-evolution between the Asian wasp and honeybee, respectively

Disclosure of Maternal HIV Status to Children: To Tell or Not To Tell . . . That Is the Question

HIV-infected mothers face the challenging decision of whether to disclose their serostatus to their children. From the perspective of both mother and child, we explored the process of disclosure, providing descriptive information and examining the relationships among disclosure, demographic variables, and child adjustment. Participants were 23 mothers and one of their noninfected children (9 to 16 years of age). Sixty-one percent of mothers disclosed. Consistent with previous research, disclosure was not related to child functioning. However, children sworn to secrecy demonstrated lower social competence and more externalizing problems. Differential disclosure, which occurred in one-third of the families, was associated with higher levels of depressive and anxiety symptoms. Finally, knowing more than mothers had themselves disclosed was related to child maladjustment across multiple domains. Clinical implications and the need for future research are considered

Do Positive Psychological Characteristics Modify the Associations of Physical Performance With Functional Decline and Institutionalization? Findings From the Longitudinal Aging Study Amsterdam

Objectives. To investigate whether 3 positive psychological characteristics, related to sense of control, modify the associations of physical performance levels with subsequent functional decline and institutionalization.Method. One thousand five hundred and thirty-two men and women participating in the Longitudinal Aging Study Amsterdam and not living in an institution in 2005-2006 were included. Mastery, self-efficacy, investment in independence, and objective physical performance scores were ascertained in 2005-2006. Functional decline and institutionalization were assessed after 3 years of follow-up.Results. The association between lower physical performance levels and increased odds of functional decline was modified by investment in independence, with a weaker association found among people with higher investment in independence scores than in people with lower scores even after adjustment for covariates. The association between lower physical performance levels and higher odds of institutionalization was marginally weaker among those people with above median levels of mastery (test of interaction p = .08). In men, an association between general self-efficacy and functional decline was found and maintained after adjustments.Conclusions. Positive psychological characteristics, related to sense of control, play a role in the transition between stages in the disablement process. Specific psychological characteristics may be associated with different stages of the disablement process and may in turn be affected by disablement

A Rapid and Economic In-House DNA Purification Method Using Glass Syringe Filters

Background Purity, yield, speed and cost are important considerations in plasmid purification, but it is difficult to achieve all of these at the same time. Currently, there are many protocols and kits for DNA purification, however none maximize all four considerations. Methodology/Principal Findings We now describe a fast, efficient and economic in-house protocol for plasmid preparation using glass syringe filters. Plasmid yield and quality as determined by enzyme digestion and transfection efficiency were equivalent to the expensive commercial kits. Importantly, the time required for purification was much less than that required using a commercial kit. Conclusions/Significance This method provides DNA yield and quality similar to that obtained with commercial kits, but is more rapid and less costly.This research was supported by Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer reviewe

Reliability of orthostatic beat-to-beat blood pressure tests: implications for population and clinical studies

Objective: To assess the test–retest reliability of orthostatic beat-to-beat blood pressure responses to active standing and related clinical definitions of orthostatic hypotension. Methods: A random sample of community-dwelling older adults from the pan-European Survey of Health, Ageing and Retirement in Europe, Ireland underwent a health assessment that mimicked that of the Irish Longitudinal Study on Ageing. An active stand test was performed using continuous blood pressure measurements. Participants attended a repeat assessment 4–12 weeks after the initial measurement. A mixed-effects regression model estimated the reliability and minimum detectable change while controlling for fixed observer and time of day effects. Results: A total of 125 individuals underwent repeat assessment (mean age 66.2 ± 7.5 years; 55.6% female). Mean time between visits was 84.3 ± 23.3 days. There was no significant mean difference in heart rate or blood pressure recovery variables between the first and repeat assessments. Minimum detectable change was noted for changes from resting values in systolic blood pressure (26.4 mmHg) and diastolic blood pressure (13.7 mmHg) at 110 s and for changes in heart rate (10.9 bpm) from resting values at 30 s after standing. Intra-class correlation values ranged from 0.47 for nadir values to 0.80 for heart rate and systolic blood pressure values measured 110 s after standing. Conclusion: Continuous orthostatic beat-to-beat blood pressure and related clinical definitions show low to moderate reliability and substantial natural variation over a 4–12-week period. Understanding variation in measures is essential for study design or estimating the effects of orthostatic hypotension, while clinically it can be used when evaluating longer term treatment effects