bluuebunny/crossref_metadata_2025_split huggingface dataset:
Labeling Tetracysteine-Tagged Proteins with Biarsenical Dyes for Live Cell Imaging
Authors: Gaietta Guido M. , Deerinck Thomas J. , Ellisman Mark H. 
Published: January 2011

INTRODUCTIONCorrelation of real-time or time-lapse light microscopy (LM) with electron microscopy (EM) of cells can be performed with biarsenical dyes. These dyes fluorescently label tetracysteine-tagged proteins so that they can be imaged with LM and, upon fluorescent photoconversion of 3,3′-diaminobenzidine tetrahydrochloride (DAB), with EM as well. In the following protocol, cells expressing tetracysteine-tagged proteins are labeled for 1 h with biarsenical dyes. The volumes indicated are for a single 30-mm culture dish containing 2 mL of labeling medium. Scale the suggested volumes up or down depending upon the size of the culture dish used in the labeling. The same procedure can be adapted for longer labeling times by lowering the amount of dye used to 50-100 nM; however, the amount of the competing dithiol EDT is maintained at 10-20 μM. Longer labeling times often produce higher signal-to-noise ratios and cause less trauma to the treated cells prior to imaging.
Fluorescence Photoconversion of Biarsenical-Labeled Cells for Correlated Electron Microscopy (EM)
Authors: Gaietta Guido M. , Deerinck Thomas J. , Ellisman Mark H. 
Published: January 2011

INTRODUCTIONCorrelated light microscopy (LM)/electron microscopy (EM) analysis can be achieved by using biarsenical dyes to fluorescently label tetracysteine-tagged proteins. Once live cell imaging using LM is complete, cellular activity can be halted promptly using a glutaraldehyde-based fixative. Rapid fixation preserves cellular ultrastructure and limits diffusion of reaction products. This protocol provides details on rapid fixation of cells, followed by fluorescence photoconversion of 3,3′-diaminobenzidine tetrahydrochloride (DAB) and sample processing for EM that can be correlated with the live cell LM images.
Preparation of Dissociated Ommatidia from Drosophila
Authors: Dolph Patrick , Nair Amit , Raghu Padinjat 
Published: January 2011

INTRODUCTIONAdult Drosophila have a compound eye composed of approximately 750 repeating units, called ommatidia, packed together to form the retina. Each ommatidium is a precise arrangement of 19 cells: eight photoreceptors (primary sensory neurons) and 11 accessory cells. Analysis of visual physiology in Drosophila photoreceptors has been central to understanding a number of important areas of modern biology, including the G-protein-coupled receptor cycle, phosphoinositide signaling, and calcium signaling. Analysis of photoreceptor performance and synaptic transmission are areas of neurobiology that have been studied using Drosophila photoreceptors as a model system. Electrophysiological analysis of responses to light is a powerful tool for characterizing and understanding visual transduction in Drosophila photoreceptors. This protocol describes the isolation of dissociated ommatidia for use in voltage-clamp recordings of currents from photoreceptors, allowing detailed analysis of ion channels such as the transient receptor potential and potassium channels in photoreceptors. Mechanical dissociation of ommatidia is typically performed in flies that are <4 h post-eclosion. In animals older than this, the ommatidia become progressively more firmly attached to each other and cannot be dissociated mechanically without causing substantial damage to the constituent photoreceptors. Like the rest of the fly, the eye is surrounded by a cuticle. Thus, before the dissociation process, the eye is dissected from the rest of the head and the retina is scooped out of the cuticular covering.
Mounting Live Cells onto Microscope Slides
Authors: Chazotte Brad 
Published: January 2011

INTRODUCTIONThis protocol describes the mounting of cells which have been grown on cleaned coverslips in cell culture dishes containing the appropriate medium and supplements. Labeling is performed with the coverslips placed in culture dishes. After cells are labeled, the coverslips can be mounted onto microscope slides to be viewed for short-term observation of live cells or longer-term observation of fixed cells. For viewing live cells for longer periods of time, specialized chambers should be used in which suitable growth conditions can be maintained.
Labeling Nuclear DNA Using DAPI
Authors: Chazotte Brad 
Published: January 2011

INTRODUCTIONA number of fluorescent stains are available that label DNA and allow easy visualization of the nucleus in interphase cells and chromosomes in mitotic cells, including Hoechst, 4′,6-diamidino-2-phenylindole (DAPI), ethidium bromide, propidium iodide, and acridine orange. Although not as bright as the vital Hoechst stains for DNA, DAPI has greater photostability. It is believed that DAPI associates with the minor groove of double-stranded DNA, with a preference for the adenine-thymine clusters. Cells must be permeabilized and/or fixed for DAPI to enter the cell and to bind DNA. Fluorescence increases approximately 20-fold when DAPI is bound to double-stranded DNA. This protocol describes the use of DAPI to label nuclear DNA of cells grown in culture.
Labeling Nuclear DNA with Hoechst 33342
Authors: Chazotte Brad 
Published: January 2011

INTRODUCTIONA number of fluorescent stains are available that label DNA and allow easy visualization of the nucleus in interphase cells and chromosomes in mitotic cells. One advantage of Hoechst 33342 is that it is membrane permeant and, thus, can stain live cells. Hoechst 33342 binds to adenine-thymine-rich regions of DNA in the minor groove. On binding to DNA, the fluorescence greatly increases. This protocol describes the use of Hoechst 33342 to label nuclear DNA of cells grown in culture.
Correlated Live Cell Light and Electron Microscopy Using Tetracysteine Tags and Biarsenicals
Authors: Gaietta Guido M. , Deerinck Thomas J. , Ellisman Mark H. 
Published: January 2011

INTRODUCTIONFundamental to obtaining a more complete understanding of the roles played by macromolecular complexes in cells is the ability to map their location, movement, and transient interactions at high temporal and high spatial resolution. Unfortunately, probes capable of allowing direct correlation of real-time or time-lapse light microscopy (LM) with electron microscopic observations are relatively few. Genetically encoded fluorescent reporters such as green fluorescent protein (GFP) have revolutionized live cell imaging studies but are not directly visible by electron microscopy (EM). Fluorescent nanoparticles or quantum dots are a type of label for LM that can also be visualized directly by EM, but targeting these to cytoplasmic proteins in living cells remains difficult. One method that does allow for highly correlated LM and EM with excellent preservation of cellular ultrastructure is fluorescence photoconversion, in which a fluorescent compound causes the deposition of a reaction product that can be rendered electron-dense and directly visualized by EM. We have used this method in combination with a class of genetically encoded peptide tags that can be labeled in living cells by fluorophores bearing two appropriately spaced arsenic atoms (biarsenicals). The tetracysteine motif is short, easily inserted into or attached to the termini of the host protein, and can be used in combination with other molecular tags such as GFP and its derivatives. This article presents methods to label cells with biarsenicals, conduct live cell imaging recording sessions, and generate specimens that can be evaluated by EM for a correlated LM/EM analysis.
Combining Enhanced Root and Shoot Growth Reveals Cross Talk between Pathways That Control Plant Organ Size in Arabidopsis
Authors: Vercruyssen Liesbeth , Gonzalez Nathalie , Werner Tomá¡ , Schmülling Thomas , Inzé Dirk 
Published: January 2011

Abstract
Functionally distinct Arabidopsis (Arabidopsis thaliana) genes that positively affect root or shoot growth when ectopically expressed were combined to explore the feasibility of enhanced biomass production. Enhanced root growth resulting from cytokinin deficiency was obtained by overexpressing CYTOKININ OXIDASE/DEHYDROGENASE3 (CKX3) under the control of the root-specific PYK10 promoter. Plants harboring the PYK10-CKX3 construct were crossed with four different transgenic lines showing enhanced leaf growth. For all combinations, the phenotypic traits of the individual lines could be combined, resulting in an overall growth increase. Unexpectedly, three out of four combinations had more than additive effects. Both leaf and root growth were synergistically enhanced in plants ectopically expressing CKX3 and BRASSINOSTEROID INSENSITIVE1, indicating cross talk between cytokinins and brassinosteroids. In agreement, treatment of PYK10-CKX3 plants with brassinolide resulted in a dramatic increase in lateral root growth that could not be observed in wild-type plants. Coexpression of CKX3 and the GROWTH-REGULATING FACTOR5 (GRF5) antagonized the effects of GRF5 overexpression, revealing an interplay between cytokinins and GRF5 during leaf cell proliferation. The combined overexpression of CKX3 and GIBBERELLIN 20-OXIDASE1 led to a synergistic increase in leaf growth, suggesting an antagonistic growth control by cytokinins and gibberellins. Only additive effects on root and shoot growth were visible in plants ectopically expressing both CKX3 and ARABIDOPSIS VACUOLAR PYROPHOSPHATASE1, hinting at an independent action mode. Our results show new interactions and contribute to the molecular and physiological understanding of biomass production at the whole plant level.
Interdecadal declines in flood frequency increase primary production in lakes of a northern river delta
Authors: McGOWAN SUZANNE , LEAVITT PETER R. , HALL ROLAND I. , WOLFE BRENT B. , EDWARDS THOMAS W. D. , KARST‐RIDDOCH TAMMY , VARDY SHEILA R. 
Published: January 2011

AbstractHuman activities and climate change have greatly altered flooding regimes in many of the world's river deltas, but the impact of such changes remains poorly quantified on decadal to multidecadal timescales. This study identified the response of delta lake primary production (measured as the concentration of sedimentary pigments) to variations in flood frequency using spatial surveys and paleolimnological analyses of lakes in the Peace‐Athabasca Delta (PAD), Canada. Surveys of 61 lakes spanning a range of hydrological conditions showed that those lakes that received flood waters less frequently were associated with elevated algal production (surface sedimentary pigments) and, in some lakes, increased growth of emergent macrophytes and epiphytic diatoms. Paleolimnological analyses of five lakes corroborated the contemporary spatial survey results by showing that production of pigments from most algal groups increased during recent periods of lower flood frequency in the 20th century as determined from increases in cellulose‐inferred lake‐water oxygen isotope composition and plant macrofossils, but remained stable in a ‘reference’ basin. In general, past periods of elevated algal production coincided with the increased abundance of submerged macrophytes or emergent vegetation that provide habitat for attached algae. These results suggest that interdecadal declines in river discharge arising from increased aridity, hydrologic regulation or consumptive water use will cause long‐term increases in primary production and alter ecosystem processes (carbon sequestration, biological diversity) in aquatic delta ecosystems similar to the PAD where lakes become nutrient‐rich in the absence of flooding.
Training emotional intelligence related to treatment skills of staff working with clients with intellectual disabilities and challenging behaviour
Authors: Zijlmans L. J. M. , Embregts P. J. C. M. , Gerits L. , Bosman A. M. T. , Derksen J. J. L. 
Published: January 2011

AbstractBackground  Staff working with clients with intellectual disabilities (ID) who display challenging behaviour may contribute to the continuation of this behaviour, because it causes emotional reactions such as anxiety, anger and annoyance, which may prohibit adequate response behaviour. To enhance staff behaviour and treatment skills a training that aimed at improving emotional intelligence (EQ) was developed.Aim  The goal of this study was to assess whether an EQ training in combination with a video‐feedback training programme improves emotional intelligence of staff working with clients with ID and challenging behaviour.Methods  Participants were 60 staff members working with individuals with ID and challenging behaviour. Thirty‐four staff members participated in a 4‐month training programme and 26 constituted the control group. A pretest–posttest control group design was used. Effectiveness was assessed by using the Dutch version of the Bar‐On EQ‐i and the judgments of experts on emotional intelligence.Results  Emotional intelligence of the experimental group changed significantly more than that of the control group. Judgments of experts on emotional intelligence indicated that the change of emotional intelligence of the experimental group improved positively.Conclusions  The positive effect of the training programme on emotional intelligence is consistent with previous research on emotional intelligence and suggests that emotional intelligence of staff working with clients with ID and challenging behaviour can be influenced by training.
The relationship between specific cognitive impairment and behaviour in Prader–Willi syndrome
Authors: Woodcock K. A. , Oliver C. , Humphreys G. W. 
Published: January 2011

AbstractBackground  Individuals with Prader–Willi syndrome (PWS) have been shown to demonstrate a particular cognitive deficit in attention switching and high levels of preference for routine and temper outbursts. This study assesses whether a specific pathway between a cognitive deficit and behaviour via environmental interaction can exist in individuals with PWS.Methods  Four individuals with PWS participated in a series of three single‐case experiments including laboratory‐based and natural environment designs. Cognitive (computer‐based) challenges placed varying demands on attention switching or controlled for the cognitive demands of the tasks while placing no demands on switching. Unexpected changes to routines or expectations were presented in controlled games, or imposed on participants' natural environments and compared with control conditions during which no unexpected changes occurred. Behaviour was observed and heart rate was measured.Results  Participants showed significantly increased temper outburst related behaviours during cognitive challenges that placed demands on attention switching, relative to the control cognitive challenges. Participants showed significantly increased temper outburst related behaviours when unexpected changes occurred in an experimental or the natural environment compared with when no changes occurred.Conclusions  Difficult behaviours that could be triggered reliably in an individual by a specific cognitive demand could also be triggered via manipulation of the environment. Results suggest that a directional relationship between a specific cognitive deficit and behaviour, via environmental interaction, can exist in individuals with PWS.
A pilot study of social competence group training for adolescents with borderline intellectual functioning and emotional and behavioural problems (SCT‐ABI)
Authors: Nestler J. , Goldbeck L. 
Published: January 2011

AbstractBackground  Emotional and behavioural problems as well as a lack of social competence are common in adolescents with borderline intellectual functioning and impair their social and vocational integration. Group interventions specifically developed for this target group are scarce and controlled evaluation studies are absent.Methods  A cognitive–behavioural group training (Social Competence Training for Adolescents with Borderline Intelligence) was developed and its effectiveness was examined in students attending special vocational schools. A total of 77 adolescents with borderline intelligence were randomised either to the intervention (n = 40) or to the control group (n = 37). Outcome measures at post‐treatment and at a 6‐month follow‐up comprised self‐reports, caregiver reports and behavioural observations.Results  The adolescents in the intervention group showed temporally stable improvement in their social competence, especially in social problem solving (F = 17.6, P < 0.001) and attainment of individual behavioural goals in everyday life (self‐reports: F = 15.9, P < 0.001; caregiver reports: F = 87.9, P < 0.001). Effects of the intervention on other outcomes such as problem‐solving competence and skills performed in standardised role plays were weak or absent.Conclusions  Social Competence Training for Adolescents with Borderline Intelligence is a promising treatment for adolescents with borderline intellectual functioning and associated emotional/behavioural symptoms. Further studies examining long‐term effects on the participants’ social and vocational integration are needed.
Stereotyped behaviour in children with autism and intellectual disability: an examination of the executive dysfunction hypothesis
Authors: Sayers N. , Oliver C. , Ruddick L. , Wallis B. 
Published: January 2011

AbstractBackground  Increasing attention has been paid to the executive dysfunction hypothesis argued to underpin stereotyped behaviour in autism. The aim of this study is to investigate one component of this model, that stereotyped behaviours are related to impaired generativity and compromised behavioural inhibition, by examining whether episodes of these behaviours terminate naturally without external intervention.Methods  Using a naturalistic observational methodology the stereotyped behaviours of six participants with autism were recorded in real time over periods ranging from 3.59 to 9.20 h. Data were also recorded for teaching staff interactions with participants and environmental settings (one‐to‐one, group and freetime).Results  In comparison with one‐to‐one settings stereotyped behaviours were more frequent when participants were in freetime settings with the exception of one participant. For the termination of stereotyped behaviours these behaviours stopped with no ongoing physical or verbal adult contact being evident for a median of 73.25% of the time. Additionally, for a median of 89.40% of the time stereotyped behaviour stopped without the initiation of adult contact.Discussion  The termination of bouts of stereotyped behaviour was frequently not associated with any adult contact and thus this form of external intervention. This finding warrants explanation by extending the hypothesis that compromised impaired generativity and behavioural inhibition offers a complete explanation of stereotyped behaviours.
Prevalence, phenomenology, aetiology and predictors of challenging behaviour in Smith‐Magenis syndrome
Authors: Sloneem J. , Oliver C. , Udwin O. , Woodcock K. A. 
Published: January 2011

AbstractBackground The prevalence, phenomenology aetiology and correlates of four forms of challenging behaviour in 32 children and adults with Smith‐Magenis syndrome (SMS) were investigated.Methods Cognitive assessments, questionnaires and semi‐structured interviews were used to gather data on intellectual disability, verbal and physical aggression, destructive behaviour and self‐injury and on characteristics known to be associated with aggression.Results Aggression in SMS was more prevalent (87%), but not more severe than aggression in contrast groups. Aggressive behaviour was more frequently associated with environmental contingencies (e.g. attention, escape and access to tangibles) than self‐injury and destructive behaviours. Severity of challenging behaviours was associated with high impulsivity.Conclusion Aggression is seen in the majority of people with SMS. Results suggest that behavioural disinhibition and operant social reinforcement are associated with the manifestation of aggression.
Association between selected folate pathway polymorphisms and nonsyndromic limb reduction defects: a case–parental analysis
Authors: Cleves Mario A. , Hobbs Charlotte A. , Zhao Weizhi , Krakowiak Patrycja A. , MacLeod Stewart L. ,   the National Birth Defects Prevention Study
Published: January 2011

SummaryCleves MA, Hobbs CA, Zhao W, Krakowiak PA, MacLeod SL, the National Birth Defects Prevention Study. Association between selected folate pathway polymorphisms and nonsyndromic limb reduction defects: a case–parental analysis. Paediatric and Perinatal Epidemiology 2011.Inadequate folate status resulting from either genetic variation or nutritional deficiencies has been associated with an increased risk of congenital malformations including orofacial clefting, limb, cardiac and neural tube defects. Few epidemiological studies have examined the association between limb reduction defects (LRDs) and folate‐related genetic polymorphisms other than MTHFR 677C→T. We conducted a case–parental analysis of 148 families who participated in the National Birth Defects Prevention Study to examine the association between nonsyndromic transverse and longitudinal LRDs with five single nucleotide polymorphisms (SNPs) in genes encoding enzymes in folate and methionine pathways. Log‐linear Poisson regression, adapted for analysis of case–parental data assuming an additive genetic model, was used to estimate genetic relative risks and 95% confidence intervals for the association between LRDs and each SNP. Among women who did not take multivitamin supplements, the MTHFR 677T variant acts via the offspring's genome to increase the risk of LRDs. No association between LRDs and any fetal SNP was found among women who used multivitamin supplements. These results suggest the possibility that initiating folic acid supplementation prior to pregnancy may reduce the risk of having a LRD‐affected pregnancy, especially in women whose offspring inherit one or two copies of the MTHFR 677T variant.
Age at diagnosis and disease progression of cystic fibrosis in an area without newborn screening
Authors: de Monestrol Isabelle , Klint Åsa , Sparén Pär , Hjelte Lena 
Published: January 2011

Summaryde Monestrol I, Klint Å, Sparén P, Hjelte L. Age at diagnosis and disease progression of cystic fibrosis in an area without newborn screening. Paediatric and Perinatal Epidemiology 2011; 25: 298–305.We studied age at diagnosis and disease progression of cystic fibrosis (CF) patients with a new study design, using data of 119 patients extracted from Stockholm CF Centre registry. Risk factors for overall morbidity and for lung, liver and nutritional morbidity were investigated separately using time to event methodology (Kaplan‐Meier curves, proportional hazards regression). The patients were followed from: (i) healthy at diagnosis to morbidity, (ii) diagnosis with symptoms of morbidity to being free of morbidity, and (iii) free of morbidity to relapse of morbidity.Median age at diagnosis was 5.0 months. Of the patients with overall morbidity at diagnosis 50% became free of morbidity after 4.8 years; however, the patients above the age of 24 months at diagnosis had a reduced chance of becoming free of morbidity (crude hazard ratio 0.14 [95 % confidence interval 0.04, 0.45]) compared with those with diagnosis between the ages of 2 and 12 months (P < 0.01).Of the healthy at diagnosis, 50% experienced overall morbidity after 1.4 years. They had a slow decline to the endpoint of the specific morbidities; 50% experienced lung morbidity after 3.4 years and liver morbidity after 4.8 years, while 50% never reached nutritional morbidity during the 10 years follow‐up.We conclude that there was a disadvantage for the CF patients diagnosed after the age of 24 months with symptoms of overall morbidity at diagnosis in an area without newborn screening.