Use of genetic decompositions to scaffold the development of a structurally sequenced curriculum for mathematics acceleration

The authors have collaboratively used a graphical language to describe their shared knowledge of a small domain of mathematics, which has in turn scaffolded their re-development of a related curriculum for mathematics acceleration. This collaborative use of the graphical language is reported as a simple descriptive case study. This leads to an evaluation of the graphical language’s usefulness as a tool to support the articulation of the structure of mathematics knowledge. In turn, implications are drawn for how the graphical language may be utilised as the detail of the curriculum is further elaborated and communicated to teachers.

Binding properties of peptidic affinity ligands for plasmid DNA capture and detection

Peptides constructed from α-helical subunits of the Lac repressor protein (LacI) were designed then tailored to achieve particular binding kinetics and dissociation constants for plasmid DNA purification and detection. Surface plasmon resonance was employed for quantification and characterization of the binding of double stranded Escherichia coli plasmid DNA (pUC19) via the lac operon (lacO) to "biomimics" of the DNA binding domain of LacI. Equilibrium dissociation constants (K D), association (k a), and dissociation rates (k d) for the interaction between a suite of peptide sequences and pUC19 were determined. K D values measured for the binding of pUC19 to the 47mer, 27mer, 16mer, and 14mer peptides were 8.8 ± 1.3 × 10 -10 M, 7.2 ± 0.6 × 10 -10 M, 4.5 ± 0.5 × 10 -8 M, and 6.2 ± 0.9 × 10 -6 M, respectively. These findings show that affinity peptides, composed of subunits from a naturally occurring operon-repressor interaction, can be designed to achieve binding characteristics suitable for affinity chromatography and biosensor devices.

Binding mechanism of affinity ligands for purification of plasmid DNA

Plasmid DNA for therapeutic and vaccination purposes must be highly purified. The high selectivity of affinity chromatography makes it ideal for the isolation of pDNA from complex biological feed stocks. Affinity chromatography makes use of the biological function and/or individual chemical structure of the interacting molecules. However, the success of any affinity purification protocol is dependent on the availability of suitable ligands. In this study, surface plasmon resonance (SPR) based Biacore system has been employed for the detection and quantification of the binding between lac operon (lacO) sequence contained in a pDNA and synthetic peptides based on the DNA-binding domain of the lac repressor protein, lad. The equilibrium dissociation constant (K D) and association and dissociation rate constants (ka, kd) for the interaction between plasmid DNA and designed peptides were determined.

LacO-LacI interaction in affinity adsorption of plasmid DNA

Current approaches for purifying plasmids from bacterial production systems exploit the physiochemical properties of nucleic acids in non-specific capture systems. In this study, an affinity system for plasmid DNA (pDNA) purification has been developed utilizing the interaction between the lac operon (lacO) sequence contained in the pDNA and a 64mer synthetic peptide representing the DNA-binding domain of the lac repressor protein, LacI. Two plasmids were evaluated, the native pUC19 and pUC19 with dual lacO3/lacOs operators (pUC19lacO3/lacOs), where the lacOs operator is perfectly symmetrical. The DNA-protein affinity interaction was evaluated by surface plasmon resonance using a Biacore system. The affinity capture of DNA in a chromatography system was evaluated using LacI peptide that had been immobilized to Streamline™ adsorbent. The KD-values for double stranded DNA (dsDNA) fragments containing lacO1 and lacO3 and lacOs and lacO3 were 5.7 ± 0.3 × 10 -11 M and 4.1 ± 0.2 × 10-11 M respectively, which compare favorably with literature reports of 5 × 10-10 - 1 × 10-9 M for native laCO1 and 1-1.2 × 10-10 M for lacO1 in a saline buffer. Densitometric analysis of the gel bands from the affinity chromatography run clearly showed a significant preference for capture of the supercoiled fraction from the feed pDNA sample. The results indicate the feasibility of the affinity approach for pDNA capture and purification using native protein-DNA interaction.

A new period of volcanogenic massive sulfide formation in the Yilgarn: A volcanological study of the ca 2.76 Ga Hollandaire VMS deposit, Yilgarn Craton, Western Australia

The Archean Hollandaire volcanogenic massive sulfide deposit is a felsic–siliciclastic VMS deposit located in the Murchison Domain of the Youanmi Terrane, Yilgarn Craton, Western Australia. It is hosted in a succession of turbidites, mudstones and coherent rhyodacite sills and has been metamorphosed to upper greenschist/lower amphibolite facies and includes a pervasive S1 deformational fabric. The coherent rhyodacitic sills are interpreted as syndepositional based on geochemical similarities with well-known VMS-associated felsic rocks and similar foliations to the metasediments. We offer several explanations for the absence of textural evidence (e.g. breccias) for syn-depositional origins: 1) the subaqueous sediments were dehydrated by long-lived magmatism such that no pore-water remained to drive quench fragmentation; 2) pore-space occlusion by burial and/or, 3) alteration overprinting and obscuring of primary breccias at contact margins. Mineralisation occurs by sub-seafloor replacement of original host rocks in two ore bodies, Hollandaire Main (~125 x >500 m and ~8 m thick) and Hollandaire West (~100 x 470 m and ~5 m thick), and occurs in three main textural styles, massive sulfides, which are exclusively hosted in turbidites and mudstones, and stringer and disseminated sulfides, which are also hosted in coherent rhyodacite. Most sulfides have textures consistent with remobilisation and recrystallisation. Hydrothermal metamorphism has altered the hangingwall and footwall to similar degrees, with significant gains in Mg, Mn and K and losses in Na, Ca and Sr. Garnet and staurolite porphyryoblasts also exhibit a footprint around mineralisation, extending up to 30 m both above and below the ore zone. High precision thermal ionisation mass spectrometry of zircons extracted from the coherent rhyodacite yield an age of 2759.5 ± 0.9 Ma, which along with geochemical comparisons, places the succession within the 2760–2735 Ma Greensleeves Formation of the Polelle Group of the Murchison Supergroup. Geochemical and geochronological evidence link the coherent rhyodacite sills to the Peter Well Granodiorite pluton ~2 km to the W, which acted as the heat engine driving hydrothermal circulation during VMS mineralisation. This study highlights the importance of both: detailed physical volcanological studies from which an accurate assessment of timing relationships, particularly the possibility of intrusions dismembering ore horizons, can be made; and identifying synvolcanic plutons and other similar suites, for VMS exploration targets in the Youanmi Terrane and worldwide.

Analysing governance of Australia's system of landscape-based greenhouse gas abatement

Healthy governance systems are key to delivering effective outcomes in any broad domain of natural resource management (NRM). One of Australia's emerging NRM governance domains is our national framework for greenhouse gas abatement (GGA), as delivered through a wide range of management practices in the Australian landscape. The emerging Landscape-Based GGA Domain represents an innovative governance space that straddles both the nation's broader NRM Policy and Delivery Domain and Australia's GGA Domain. As a point-in-time benchmark, we assess the health of this hybrid domain as it stood at the end of 2013. At that time, the domain was being progressed through the Australian government's Clean Energy Package and, more particularly, its Carbon Farming Initiative (CFI). While significant changes are currently under development by a new Australian government, this paper explores key areas of risk within the governance system underpinning this emerging hybrid domain at that point in time. We then map some potential reform or continuous improvement pathways required (from national to paddock scale) with the view to securing improved landscape outcomes over time through widespread GGA activities.

The quality of life and caregiving burden among caregivers of people with dementia in Hanoi, Bac Ninh and Hai Phong, Vietnam

This three phase study aimed to describe dementia carer's quality of life (QoL) and perceived burden, and explore the associations between family carer characteristics, burden and perceived QoL in Vietnam. Dementia carers in the capital, Hanoi, in Phase 1 (N= 153) and from Hanoi, Hai Phong and Bac Ninh in Phase 2 (N=347) completed questionnaires. Survey results showed dementia carers reported low QoL, predicted by high perceived burden. Other carer characteristics including age, gender, family income and perceived experience were significantly associated with QoL. Filial piety contributed to only a single domain of QoL. Phase 3 employed qualitative methods to explore the specific issues faced by daughter carers. Findings suggested that filial gratitude and positive aspects of the role may influence the caring experience of daughter carers. Further investigation of the specific support needs of general dementia carers, and daughter carers in particular, in Vietnam is warranted.

EphB4 localises to the nucleus of prostate cancer cells

The EphB4 receptor tyrosine kinase is over-expressed in a variety of different epithelial cancers including prostate where it has been shown to be involved in survival, migration and angiogenesis. We report here that EphB4 also resides in the nucleus of prostate cancer cell lines. We used in silico methods to identify a bipartite nuclear localisation signal (NLS) in the extracellular domain and a monopartite NLS sequence in the intracellular kinase domain of EphB4. To determine whether both putative NLS sequences were functional, fragments of the EphB4 sequence containing each NLS were cloned to create EphB4NLS-GFP fusion proteins. Localisation of both NLS-GFP proteins to the nuclei of transfected cells was observed, demonstrating that EphB4 contains two functional NLS sequences. Mutation of the key amino residues in both NLS sequences resulted in diminished nuclear accumulation. As nuclear translocation is often dependent on importins we confirmed that EphB4 and importin-α can interact. To assess if nuclear EphB4 could be implicated in gene regulatory functions potential EphB4-binding genomic loci were identified using chromatin immunoprecipitation and Lef1 was confirmed as a potential target of EphB4-mediated gene regulation. These novel findings add further complexity to the biology of this important cancer-associated receptor.

Household carbon emission research: An analytical review of measurement, influencing factors and mitigation prospects

Greenhouse gas (GHG) emissions are simultaneously exhausting the world's supply of fossil fuels and threatening the global climate. In many developing countries, significant improvement in living standards in recent years due to the accelerating development of their economies has resulted in a disproportionate increase in household energy consumption. Therefore, a major reduction in household carbon emissions (HCEs) is essential if global carbon reduction targets are to be met. To do this, major Organisation for Economic Co-operation and Development (OECD) states have already implemented policies to alleviate the negative environmental effects of household behaviors and less carbon-intensive technologies are also proposed to promote energy efficiency and reduce carbon emissions. However, before any further remedial actions can be contemplated, though, it is important to fully understand the actual causes of such large HCEs and help researchers both gain deep insights into the development of the research domain and identify valuable research topics for future study. This paper reviews existing literature focusing on the domain of HCEs. This critical review provides a systematic understanding of current work in the field, describing the factors influencing HCEs under the themes of household income, household size, age, education level, location, gender and rebound effects. The main quantification methodologies of input–output models, life cycle assessment and emission coefficient methods are also presented, and the proposed measures to mitigate HCEs at the policy, technology and consumer levels. Finally, the limitations of work done to date and further research directions are identified for the benefit of future studies.

VH gene usage in immunoglobulin E responses of seasonal rhinitis patients allergic to grass pollen is oligoclonal and antigen driven

Background: IgE is the pivotal-specific effector molecule of allergic reactions yet it remains unclear whether the elevated production of IgE in atopic individuals is due to superantigen activation of B cell populations, increased antibody class switching to IgE or oligoclonal allergen-driven IgE responses. Objectives: To increase our understanding of the mechanisms driving IgE responses in allergic disease we examined immunoglobulin variable regions of IgE heavy chain transcripts from three patients with seasonal rhinitis due to grass pollen allergy. Methods: Variable domain of heavy chain-epsilon constant domain 1 cDNAs were amplified from peripheral blood using a two-step semi-nested PCR, cloned and sequenced. Results: The VH gene family usage in subject A was broadly based, but there were two clusters of sequences using genes VH 3-9 and 3-11 with unusually low levels of somatic mutations, 0-3%. Subject B repeatedly used VH 1-69 and subject C repeatedly used VH 1-02, 1-46 and 5a genes. Most clones were highly mutated being only 86-95% homologous to their germline VH gene counterparts and somatic mutations were more abundant at the complementarity determining rather than framework regions. Multiple sequence alignment revealed both repeated use of particular VH genes as well as clonal relatedness among clusters of IgE transcripts. Conclusion: In contrast to previous studies we observed no preferred VH gene common to IgE transcripts of the three subjects allergic to grass pollen. Moreover, most of the VH gene characteristics of the IgE transcripts were consistent with oligoclonal antigen-driven IgE responses.

A comparative study of antibody expressions in primary biliary cirrhosis and autoimmune cholangitis using phage display

Primary biliary cirrhosis (PBC) and autoimmune cholangitis (AIC) are serologic expressions of an autoimmune liver disease affecting biliary ductular cells. Previously we screened a phage-displayed random peptide library with polyclonal IgG from 2 Australian patients with PBC and derived peptides that identified a single conformational (discontinuous) epitope in the inner lipoyl domain of the E2 subunit of the pyruvate dehydrogenase complex (PDC-E2), the characteristic autoantigen in PBC. Here we have used phage display to investigate the reactivity of PBC sera from 2 ethnically and geographically distinct populations, Japanese and Australian, and the 2 serologic expressions, PBC and AIC. Random 7-mer and 12-mer peptide libraries were biopanned with IgG from 3 Japanese patients with PBC and 3 with AIC who did not have anti-PDC-E2. The phage clones (phagotopes) obtained were tested by capture enzyme-linked immunosorbent assay (ELISA) for reactivity with affinity-purified anti-PDC-E2, and compared with those obtained from Australian patients with PBC. Peptide sequences of the derived phagotopes and sequences derived by biopanning with irrelevant antisera were aligned to develop a guide tree based on physicochemical similarity. Both Australian and Japanese PBC-derived phagotopes were distributed in branches of the guide tree that contained the peptide sequences MH and FV previously identified as part of an immunodominant conformational epitope of PDC-E2, indicating that epitope selection was not influenced by the racial origin of the PBC sera. Biopanning with either PBC or AIC-derived IgG yielded phagotopes that reacted with anti-PDC-E2 by capture ELISA, further establishing that there is a similar autoimmune targeting in PBC and AIC.

Towards the pathogenesis of autoimmune liver disease

There have been recent improvements in the clinical understanding and definition of the major types of autoimmune liver disease. However, still lacking is knowledge of their prevalence and pathogenesis. Three areas of study are in progress in our laboratory. First, in type 1 autoimmune hepatitis, the search continues to identify a liver/disease-specific autoantigenic reactant. Using hepatocyte membrane preparations, immunoblotting has underlined the problem of distinguishing, among multiple reactants, those that may be causally rather than consequentially related to hepatocellular damage. Second, in primary biliary cirrhosis (PBC), the need for population screening to ascertain prevalence and detect preclinical cases can be met by a rapid automated procedure for detection, by specific enzyme inhibition in microtitre wells, of antibody (anti-M2) to the pyruvate dehydrogenase complex E2 subunit (PDC-E2). Third, the structure of the conformational epitope within the inner lipoyl domain of PDC-E2 is being investigated by screening random phage-displayed peptide libraries using PBC sera. This has yielded phage clones in which the sequence of the peptide insert portrays the structure of this epitope, as judged by clustering of PBC-derived sequences to particular branches of a guide-tree that shows relatedness of peptides, and by reactivity of selected phage clones with anti-PDC-E2. Thus phage display identifies a peptide 'mimotope' of the antibody epitope in the inner lipoyl domain of PDC-E2.

Antibody reactivity to the transmembrane protein of the caprine arthritis encephalitis virus correlates with severity of arthritis: No evidence for the involvement of epitope mimicry

Serum and synovial antibody reactivities of caprine arthritis encephalitis virus (CAEV) infected goats were assessed by Western blotting against purified CAEV antigen and the greatest intensity of reactivity in the serum of arthritic goats was to the gp45 transmembrane protein (TM). The extracytoplasmic domain of the TM gene was cloned into a pGEX vector and expressed in Escherichia coil as a glutathione S transferase fusion protein (GST-TM). This clone was found to be 90.5 and 89.2% homologous to published sequences of CAEV TM gene. Serum of 16 goats naturally infected with CAEV were examined by Western blotting for reactivity to the fusion protein. Antibody reactivity to the GST-TM correlated with clinically detectable arthritis (R = 0.642, P ≤ 0.007). The hypothesis that the immune response to the envelope proteins of the CAEV contributes to the severity of arthritis in goats naturally infected with CAEV via epitope mimicry was tested. Antibodies from 5 CAEV infected goats were affinity purified against the GST-TM fusion protein and tested for cross-reactivity with a series of goat synovial extracts and proteogylcans. No serum antibody response or cross-reactivity of affinity purified antibodies could be detected. Peptides of the CAEV SU that were predicted to be linear epitopes and a similar heat shock protein 83 (HSP) peptide identified by database searching, were synthesized and tested for reactivity in CAEV goats using ELISA, in vitro lymphocyte proliferation and delayed type hypersensitivity (DTH) assays. Peripheral blood lymphocytes from 10 of 17 goats with long term natural CAEV infections proliferated in vitro in response to CAEV and in vivo 3 of 7 CAEV infected goats had a DTH reaction to CAEV antigen. However, none of the peptides elicited significant cell mediated immune responses from CAEV infected goats. No antibody reactivity to the SU peptides or HSP peptide was found. We observed that the antibody reactivity to the CAEV TM protein associated with severity of arthritis however epitope mimicry by the envelope proteins of CAEV is unlikely to be involved.

Evidence-based recommendations for the monitoring and treatment of ankylosing spondylitis: Results from the Australian 3E initiative in rheumatology

Aim: To develop a set of Australian recommendations for the monitoring and treatment of ankylosing spondylitis (AS) through systematic literature review combined with the opinion of practicing rheumatologists. Methods: A set of eight questions, four in each domain of monitoring and treatment, were formulated by voting and the Delphi method. The results of a systematic literature review addressing each question were presented to the 23 participants of the Australian 3E meeting. All participants were clinical rheumatologists experienced in the daily management of AS. Results: After three rounds of breakout sessions to discuss the findings of the literature review, a set of recommendations was finalized after discussion and voting. The category of evidence and strength of recommendation were determined for each proposal. The level of agreement among participants was excellent (mean 84%, range 64-100%). Conclusions: The 12 recommendations developed from evidence and expert opinion provide guidance for the daily management of AS patients. For most recommendations, we found a paucity of supportive evidence in the literature highlighting the need for additional clinical studies.

Best practice full-stop: Anticipating the spillover effects of enterprise education

This paper considers the potential flow-on benefits that may arise from the increasing provision of enterprise education. The issue of vulnerability is proposed to be a factor associated with superior learning outcomes. Therefore, this paper aims to highlight the use of student and facilitator vulnerability to increase the capacity for learning in Higher Education. It is argued that students benefit from having to struggle to find their identity within a learning environment. That group work can assist the development of confidence through which improved learning behaviours are adopted. Also, the development of good learning practices in the domain of enterprise education have the potential to aid student learning outcomes in other areas of study. This paper provides another means of valuing the potential gains to be received from enterprise education.