Plasmid DNA purification via the use of a dual affinity protein

Methods are presented for the production, affinity purification and analysis of plasmid DNA (pDNA). Batch fermentation is used for the production of the pDNA, and expanded bed chromatography, via the use of a dual affinity glutathione S-transferase (GST) fusion protein, is used for the capture and purification of the pDNA. The protein is composed of GST, which displays affinity for glutathione immobilized to a solid-phase adsorbent, fused to a zinc finger transcription factor, which displays affinity for a target 9-base pair sequence contained within the target pDNA. A Picogreen™ fluorescence assay and/or anx ethidium bromide agarose gel electrophoresis assay can be used to analyze the eluted pDNA.

Kids, counselors and troubles-telling : morality-in-action in talk on an Australian children’s helpline

This article begins with the premise that morality is an intrinsic, although often invisible, aspect of everyday social action. Drawn from a corpus of fifty audiorecorded telephone calls to Kids Helpline, an Australian helpline for children and young people, we examine one call to show how the young caller and counsellor co-construct ‘morality-in-action’. Ethnomethodological understandings and, in particular, Sacks’ (1992) description of ‘Class 2’ rules and infractions show how an adolescent caller and counsellor collaboratively assemble moral versions of the caller. In puzzling out possible motives, the caller and counsellor can be seen to be attending to the implications of different moral versions of the caller. This attribution of motives is moral work in action, with motives contingently assembled, displayed and evaluated, with such work understood as displays of moral reasoning. The counselling call makes visible the counsellor’s interactional work to support and empower the client. Analysis such as this offers counsellors ways of understanding and making visible their interactional and moral work within helpline call interactions.

Sectional analysis of stormwater treatment performance of a constructed wetland

Constructed wetlands are among the most common Water Sensitive Urban Design (WSUD) measures for stormwater treatment. These systems have been extensively studied to understand their performance and influential treatment processes. Unfortunately, most past studies have been undertaken considering a wetland system as a lumped system with a primary focus on the reduction of the event mean concentration (EMC) values of specific pollutant species or total pollutant load removal. This research study adopted an innovative approach by partitioning the inflow runoff hydrograph and then investigating treatment performance in each partition and their relationships with a range of hydraulic factors. The study outcomes confirmed that influenced by rainfall characteristics, the constructed wetland displays different treatment characteristics for the initial and later sectors of the runoff hydrograph. The treatment of small rainfall events (<15 mm) is comparatively better at the beginning of runoff events while the trends in pollutant load reductions for large rainfall events (>15 mm) are generally lower at the beginning and gradually increase towards the end of rainfall events. This highlights the importance of ensuring that the inflow into a constructed wetland has low turbulence in order to achieve consistent treatment performance for both, small and large rainfall events.

Staged homicides: An examination of common features of faked burglaries, suicides, accidents and car accidents

A staged crime scene involves deliberate alteration of evidence by the offender to simulate events that did not occur for the purpose of misleading authorities (Geberth, 2006; Turvey, 2000). This study examined 115 staged homicides from the USA to determine common elements; victim and perpetrator characteristics; and specific features of different types of staged scenes. General characteristics include: multiple victims and offenders; a previous relationship be- tween parties involved; and victims discovered in their own home, often by the offender. Staged scenes were separated by type with staged burglaries, suicides, accidents, and car accidents examined in more detail. Each type of scene displays differently with separate indicators and common features. Features of staged burglaries were: no points of entry/exit staged; non-valuables taken; scene ransacking; offender self- injury; and offenders bringing weapons to the scene. Features of staged suicides included: weapon arrangement and simulating self-injury to the victim; rearranging the body; and removing valuables. Examples of elements of staged accidents were arranging the implement/weapon and re- positioning the deceased; while staged car accidents involved: transporting the body to the vehicle and arranging both; mutilation after death; attempts to secure an alibi; and clean up at the primary crime scene. The results suggest few staging behaviors are used, despite the credibility they may have offered the façade. This is the first peer-reviewed, published study to examine the specific features of these scenes, and is the largest sample studied to date.

Investigation of fiducial marker and soft-tissue image guidance techniques in prostate radiation therapy

This thesis examines and compares imaging methods used during the radiotherapy treatment of prostate cancer. The studies found that radiation therapists were able to localise and target the prostate consistently with planar imaging techniques and that the use of small gold markers in the prostate reduced the variation in prostate localisation when using volumetric imaging. It was concluded that larger safety margins are required when using volumetric imaging without gold markers.

'I wonder what you know...': Teachers designing requests for factual information

The early years are significant in optimising children’s educational, emotional and social outcomes and have become a major international policy priority. Within Australia, policy levers have prioritised early childhood education, with a focus on program quality, as it is associated with lifelong success. Longitudinal studies have found that high quality teacher-child interactions are an essential element of high quality programs, and teacher questioning is one aspect of teacher-child interactions that has been attributed to affecting the quality of education, linking open ended questioning to higher cognitive achievement. Teachers, however, overwhelmingly ask more closed than open questions. In the classroom, like everyday interaction, questions in interaction require answers. They are used to request, offer, repair, challenge, seek agreement (Curl & Drew, 2008; Enfield, Stivers, & Levinson, 2010; Hayano, 2013; Schegloff, 2007). Teachers use questions to set agendas and manage lessons (McHoul, 1978; Mehan, 1979; Sacks, 1995), and to gauge students’ knowledge and understanding (Lerner, 1995; McHoul, 1978; Mehan, 1979). Drawing on data from the Australian Research Council project Interacting with Knowledge: Interacting with people: Web searching in early childhood, this paper focuses on an extended sequence of talk between a teacher with two students aged between 3.5 and 5 years in a preschool classroom. The episode, drawn from a corpus of over 200 hours of video recorded data, captures how the teacher and children undertake an online search for images of lady beetles and hairy caterpillars on the Web. Ethnomethodological and conversation analysis approaches examine how the teacher asks questions, which call on the children to display their factual knowledge about the search topic. The fine grained analysis shows how teachers design their interactions to prompt children’s displays of factual knowledge, and how the design of factual questions affect a student’s response in terms of what and how they respond. In focussing on how the teacher designs factual questions and how children respond to these questions it shows that question design can close down a student’s reply; or elicit a range of answers, from one word to extended more detailed responses. Understanding how the design of teachers’ questions can influence students’ responses has pedagogic implications and may support educators to make intentional decisions regarding their own questioning techniques.

Vote as you go : Blending interfaces for community engagement into the urban space

This paper presents a series of studies on situated interfaces for community engagement. Firstly, we identify five recurring design challenges as well as four common strategies used to overcome them. We then assess the effectiveness of these strategies through field studies with public polling interfaces. We developed two very different polling interfaces in the form of (1) a web application running on an iPad mounted on a stand, allowing one vote at a time, and (2) a playful full-body interaction application for a large urban screen allowing concurrent participation. We deployed both interfaces in an urban precinct with high pedestrian traffic and equipped with a large urban screen. Analysing discoverability and learnability of each scenario, we derive insights regarding effective ways of blending community engagement interfaces into the built environment, while attracting the attention of passers-by and communicating the results of civic participation.

Near infrared spectroscopy of marble from Chillagoe, Australia and in comparison with other selected natural carbonates

Marble from the Chillagoe deposits was extensively used in the construction of Australia’s parliament house. Near infrared (NIR) spectroscopy has been applied to study the quality of marble from the Chillagoe marble deposits and to distinguish between different types of marble in the Chillagoe deposits. A comparison of the NIR spectra of marble with dolomite and monohydrocalcite is made. The spectrum of the marble closely resembles that of monohydrocalcite and is different from that of dolomite. The infrared spectra of the minerals are characterised by OH and water stretching vibrations. Both the first and second fundamental overtones of these bands are observed in the NIR spectra. Marble is characterised by NIR bands at 4005, 4268 and 4340 cm–1, attributed to carbonate combination bands and overtones. Marble also shows NIR bands at 5005, 5106, 5234 and 5334 cm–1 assigned to water combination bands. Finally the NIR spectrum of the marble displays broad low-intensity features centred upon 6905 cm–1 attributed to the water first overtones. It appears feasible to identify marble through the use of NIR spectroscopy.

Projecting the new world city: The city as spectacle in an urban light festival

As a precursor to the 2014 G20 Leaders’ Summit held in Brisbane, Australia, the Queensland Government sponsored a program of G20 Cultural Celebrations, designed to showcase the Summit’s host city. The cultural program’s signature event was the Colour Me Brisbane festival, a two-week ‘citywide interactive light and projection installations’ festival that was originally slated to run from 24 October to 9 November, but which was extended due to popular demand to conclude with the G20 Summit itself on 16 November. The Colour Me Brisbane festival comprised a series projection displays that promoted visions of the city’s past, present, and future at landmark sites and iconic buildings throughout the city’s central business district and thus transformed key buildings into forms of media architecture. In some instances the media architecture installations were interactive, allowing the public to control aspects of the projections through a computer interface situated in front of the building; however, the majority of the installations were not interactive in this sense. The festival was supported by a website that included information regarding the different visual and interactive displays and links to social media to support public discussion regarding the festival (Queensland Government 2014). Festival-goers were also encouraged to follow a walking-tour map of the projection sites that would take them on a 2.5 kilometre walk from Brisbane’s cultural precinct, through the city centre, concluding at parliament house. In this paper, we investigate the Colour Me Brisbane festival and the broader G20 Cultural Celebrations as a form of strategic placemaking—designed, on the one hand, to promote Brisbane as a safe, open, and accessible city in line with the City Council’s plan to position Brisbane as a ‘New World City’ (Brisbane City Council 2014). On the other hand, it was deployed to counteract growing local concerns and tensions over the disruptive and politicised nature of the G20 Summit by engaging the public with the city prior to the heightened security and mobility restrictions of the Summit weekend. Harnessing perspectives from media architecture (Brynskov et al. 2013), urban imaginaries (Cinar & Bender 2007), and social media analysis, we take a critical approach to analysing the government-sponsored projections, which literally projected the city onto itself, and public responses to them via the official, and heavily promoted, social media hashtags (#colourmebrisbane and #g20cultural). Our critical framework extends the concepts of urban phantasmagoria and urban imaginaries into the emerging field of media architecture to scrutinise its potential for increased political and civic engagement. Walter Benjamin’s concept of phantasmagoria (Cohen 1989; Duarte, Firmino, & Crestani 2014) provides an understanding of urban space as spectacular projection, implicated in commodity and techno-culture. The concept of urban imaginaries (Cinar & Bender 2007; Kelley 2013)—that is, the ways in which citizens’ experiences of urban environments are transformed into symbolic representations through the use of imagination—similarly provides a useful framing device in thinking about the Colour Me Brisbane projections and their relation to the construction of place. Employing these critical frames enables us to examine the ways in which the installations open up the potential for multiple urban imaginaries—in the sense that they encourage civic engagement via a tangible and imaginative experience of urban space—while, at the same time, supporting a particular vision and way of experiencing the city, promoting a commodified, sanctioned form of urban imaginary. This paper aims to dissect the urban imaginaries intrinsic to the Colour Me Brisbane projections and to examine how those imaginaries were strategically deployed as place-making schemes that choreograph reflections about and engagement with the city.

Local Commons: A visual approach to collective city making through situated community engagement

Due to the numerous possibilities of voicing concerns and the flood of data we are exposed to, local issues are sometimes at risk of being overlooked. This study explores Local Commons, a design intervention in public space that combines situated digital and tangible media in order to engage communities in contributing and debating different perspectives on a given local issue. The intervention invited the community to submit images of their perspectives on the issue, which were displayed on a public screen. Via tangible buttons in front of the screen, community members then agree or disagree on the displayed perspectives, creating a space for deliberation. In a user study, we were specifically interested in testing three aspects of our intervention, which are discussed in this paper: The difference that situatedness, visual content, and tangible interaction can make to urban community engagement.

A tensor-based morphometry study of genetic influences on brain structure using a new fluid registration method

We incorporated a new Riemannian fluid registration algorithm into a general MRI analysis method called tensor-based morphometry to map the heritability of brain morphology in MR images from 23 monozygotic and 23 dizygotic twin pairs. All 92 3D scans were fluidly registered to a common template. Voxelwise Jacobian determinants were computed from the deformation fields to assess local volumetric differences across subjects. Heritability maps were computed from the intraclass correlations and their significance was assessed using voxelwise permutation tests. Lobar volume heritability was also studied using the ACE genetic model. The performance of this Riemannian algorithm was compared to a more standard fluid registration algorithm: 3D maps from both registration techniques displayed similar heritability patterns throughout the brain. Power improvements were quantified by comparing the cumulative distribution functions of the p-values generated from both competing methods. The Riemannian algorithm outperformed the standard fluid registration.

A new registration method based on Log-Euclidean tensor metrics and its application to genetic studies

In structural brain MRI, group differences or changes in brain structures can be detected using Tensor-Based Morphometry (TBM). This method consists of two steps: (1) a non-linear registration step, that aligns all of the images to a common template, and (2) a subsequent statistical analysis. The numerous registration methods that have recently been developed differ in their detection sensitivity when used for TBM, and detection power is paramount in epidemological studies or drug trials. We therefore developed a new fluid registration method that computes the mappings and performs statistics on them in a consistent way, providing a bridge between TBM registration and statistics. We used the Log-Euclidean framework to define a new regularizer that is a fluid extension of the Riemannian elasticity, which assures diffeomorphic transformations. This regularizer constrains the symmetrized Jacobian matrix, also called the deformation tensor. We applied our method to an MRI dataset from 40 fraternal and identical twins, to revealed voxelwise measures of average volumetric differences in brain structure for subjects with different degrees of genetic resemblance.

Statistically assisted fluid registration algorithm - SAFIRA

In this paper, we develop and validate a new Statistically Assisted Fluid Registration Algorithm (SAFIRA) for brain images. A non-statistical version of this algorithm was first implemented in [2] and re-formulated using Lagrangian mechanics in [3]. Here we extend this algorithm to 3D: given 3D brain images from a population, vector fields and their corresponding deformation matrices are computed in a first round of registrations using the non-statistical implementation. Covariance matrices for both the deformation matrices and the vector fields are then obtained and incorporated (separately or jointly) in the regularizing (i.e., the non-conservative Lagrangian) terms, creating four versions of the algorithm. We evaluated the accuracy of each algorithm variant using the manually labeled LPBA40 dataset, which provides us with ground truth anatomical segmentations. We also compared the power of the different algorithms using tensor-based morphometry -a technique to analyze local volumetric differences in brain structure- applied to 46 3D brain scans from healthy monozygotic twins.

A nonconservative Lagrangian framework for statistical fluid registration - SAFIRA

In this paper, we used a nonconservative Lagrangian mechanics approach to formulate a new statistical algorithm for fluid registration of 3-D brain images. This algorithm is named SAFIRA, acronym for statistically-assisted fluid image registration algorithm. A nonstatistical version of this algorithm was implemented, where the deformation was regularized by penalizing deviations from a zero rate of strain. In, the terms regularizing the deformation included the covariance of the deformation matrices Σ and the vector fields (q). Here, we used a Lagrangian framework to reformulate this algorithm, showing that the regularizing terms essentially allow nonconservative work to occur during the flow. Given 3-D brain images from a group of subjects, vector fields and their corresponding deformation matrices are computed in a first round of registrations using the nonstatistical implementation. Covariance matrices for both the deformation matrices and the vector fields are then obtained and incorporated (separately or jointly) in the nonconservative terms, creating four versions of SAFIRA. We evaluated and compared our algorithms' performance on 92 3-D brain scans from healthy monozygotic and dizygotic twins; 2-D validations are also shown for corpus callosum shapes delineated at midline in the same subjects. After preliminary tests to demonstrate each method, we compared their detection power using tensor-based morphometry (TBM), a technique to analyze local volumetric differences in brain structure. We compared the accuracy of each algorithm variant using various statistical metrics derived from the images and deformation fields. All these tests were also run with a traditional fluid method, which has been quite widely used in TBM studies. The versions incorporating vector-based empirical statistics on brain variation were consistently more accurate than their counterparts, when used for automated volumetric quantification in new brain images. This suggests the advantages of this approach for large-scale neuroimaging studies.

Mapping the regional influence of genetics on brain structure variability - A Tensor-Based Morphometry study

Genetic and environmental factors influence brain structure and function profoundly. The search for heritable anatomical features and their influencing genes would be accelerated with detailed 3D maps showing the degree to which brain morphometry is genetically determined. As part of an MRI study that will scan 1150 twins, we applied Tensor-Based Morphometry to compute morphometric differences in 23 pairs of identical twins and 23 pairs of same-sex fraternal twins (mean age: 23.8 ± 1.8 SD years). All 92 twins' 3D brain MRI scans were nonlinearly registered to a common space using a Riemannian fluid-based warping approach to compute volumetric differences across subjects. A multi-template method was used to improve volume quantification. Vector fields driving each subject's anatomy onto the common template were analyzed to create maps of local volumetric excesses and deficits relative to the standard template. Using a new structural equation modeling method, we computed the voxelwise proportion of variance in volumes attributable to additive (A) or dominant (D) genetic factors versus shared environmental (C) or unique environmental factors (E). The method was also applied to various anatomical regions of interest (ROIs). As hypothesized, the overall volumes of the brain, basal ganglia, thalamus, and each lobe were under strong genetic control; local white matter volumes were mostly controlled by common environment. After adjusting for individual differences in overall brain scale, genetic influences were still relatively high in the corpus callosum and in early-maturing brain regions such as the occipital lobes, while environmental influences were greater in frontal brain regions that have a more protracted maturational time-course.