Towards the biofortification of banana fruit for enhanced micronutrient content

In Uganda, vitamin A deficiency (VAD) and iron deficiency anaemia (IDA) are major public health problems with between 15-32% of children under 5 years of age showing VAD and 73% being anaemic. This is largely due to the fact that the staple food crop of the country, banana, is low in pro-vitamin A and iron, therefore leading to dietary deficiencies. Although worldwide progress has been made to control VAD and IDA through supplementation, food fortification and diet diversification, their long term sustainability and impact in developing countries such as Uganda is limited. The approach taken by researchers at Queensland University of Technology (QUT), Australia, in collaboration with the National Agricultural Research Organization (NARO), Uganda, to address this problem, is to generate consumer acceptable banana varieties with significantly increased levels of pro-vitamin A and iron in the fruit using genetic engineering techniques. Such an approach requires the use of suitable, well characterised genes and promoters for targeted transgene expression. Recently, a new banana phytoene synthase gene (APsy2a) involved in the synthesis of pro-vitamin A (pVA) carotenoids was isolated from a high â-carotene banana (F’ei cv Asupina). In addition, sequences of banana ferritin, an iron storage protein, have been isolated from Cavendish banana. The aim of the research described in this thesis was to evaluate the function of these genes to assess their suitability for the biofortification of banana fruit. In addition, a range of banana-derived promoters were characterised to determine their suitability for controlling the expression of transgenes in banana fruit. Due to the time constraints involved with generating transgenic banana fruit, rice was used as the model crop to investigate the functionality of the banana-derived APsy2a and ferritin genes. Using Agrobacterium-mediated transformation, rice callus was transformed with APsy2a +/- the bacterial-derived carotene desaturase gene (CrtI) each under the control of the constitutive maize poly-ubiquitin promoter (ZmUbi) or seed-specific rice glutelin1 (Gt1) promoter. The maize phytoene synthase (ZmPsy1) gene was included as a control. On selective media, with the exception of ZmUbi-CrtI-transgenic callus, all antibiotic resistant callus displayed a yellow-orange colour from which the presence of â-carotene was demonstrated using Raman spectroscopy. Although the regeneration of plants from yellow-orange callus was difficult, 16 transgenic plants were obtained and characterised from callus transformed with ZmUbi-APys2a alone. At least 50% of the T1 seeds developed a yellow-orange coloured callus which was found to contain levels of â-carotene ranging from 4.6-fold to 72-fold higher than that in non-transgenic rice callus. Using the seed-specific Gt1 promoter, 38 transgenic rice plants were generated from APsy2a-CrtI-transformed callus while 32 plants were regenerated from ZmPsy1-CrtI-transformed callus. However, when analysed for presence of transgene by PCR, all transgenic plants contained the APsy2a, ZmPsy1 or CrtI transgene, with none of the plants found to be co-transformed. Using Raman spectroscopy, no â-carotene was detected in-situ in representative T1 seeds. To investigate the potential of the banana-derived ferritin gene (BanFer1) to enhance iron content, rice callus was transformed with constitutively expressed BanFer1 using the soybean ferritin gene (SoyFer) as a control. A total of 12 and 11 callus lines independently transformed with BanFer1 and SoyFer, respectively, were multiplied and transgene expression was verified by RT-PCR. Pearl’s Prussian blue staining for in-situ detection of ferric iron showed a stronger blue colour in rice callus transformed with BanFer1 compared to SoyFer. Using flame atomic absorption spectrometry, the highest mean amount of iron quantified in callus transformed with BanFer1 was 30-fold while that obtained using the SoyFer was 14-fold higher than the controls. In addition, ~78% of BanFer1-transgenic callus lines and ~27% of SoyFer-transgenic callus lines had significantly higher iron content than the non-transformed controls. Since the genes used for enhancing micronutrient content need to be expressed in banana fruit, the activity of a range of banana-derived, potentially fruit-active promoters in banana was investigated. Using uidA (GUS) as a reporter gene, the function of the Expansin1 (MaExp1), Expansin1 containing the rice actin intron (MaExp1a), Expansin4 (MaExp4), Extensin (MaExt), ACS (MaACS), ACO (MaACO), Metallothionein (MaMT2a) and phytoene synthase (APsy2a) promoters were transiently analysed in intact banana fruit using two transformation methods, particle bombardment and Agrobacterium-mediated infiltration (agro-infiltration). Although a considerable amount of variation in promoter activity was observed both within and between experiments, similar trends were obtained using both transformation methods. The MaExp1 and MaExp1a directed high levels of GUS expression in banana fruit which were comparable to those observed from the ZmUbi and Banana bunchy top virus-derived BT4 promoters that were included as positive controls. Lower levels of promoter activity were obtained in both methods using the MaACO and MaExt promoters while the MaExp4, MaACS, and APsy2a promoters directed the lowest GUS activity in banana fruit. An attempt was subsequently made to use agro-infiltration to assess the expression of pVA biosynthesis genes in banana fruit by infiltrating fruit with constructs in which the ZmUbi promoter controlled the expression of APsy2a +/- CrtI, and with the maize phytoene synthase gene (ZmPsy1) included as a control. Unfortunately, the large amount of variation and inconsistency observed within and between experiments precluded any meaningful conclusions to be drawn. The final component of this research was to assess the level of promoter activity and specificity in non-target tissue. These analyses were done on leaves obtained from glasshouse-grown banana plants stably transformed with MaExp1, MaACO, APsy2a, BT4 and ZmUbi promoters driving the expression of the GUS gene in addition to leaves from a selection of the same transgenic plants which were growing in a field trial in North Queensland. The results from both histochemical and fluorometric GUS assays showed that the MaExp1 and MaACO promoters directed very low GUS activities in leaves of stably transformed banana plants compared to the constitutive ZmUbi and BT4 promoters. In summary, the results from this research provide evidence that the banana phytoene synthase gene (APsy2a) and the banana ferritin gene (BanFer1) are functional, since the constitutive over-expression of each of these transgenes led to increased levels of pVA carotenoids (for APsy2a) and iron content (for BanFer1) in transgenic rice callus. Further work is now required to determine the functionality of these genes in stably-transformed banana fruit. This research also demonstrated that the MaExp1 and MaACO promoters are fruit-active but have low activity in non-target tissue (leaves), characteristics that make them potentially useful for the biofortification of banana fruit. Ultimately, however, analysis of fruit from field-grown transgenic plants will be required to fully evaluate the suitability of pVA biosynthesis genes and the fruit-active promoters for fruit biofortification.

Georg Calixtus, Isaac Casaubon, and the Consensus of Antiquity

This article explores the relationship between Georg Calixtus (1586-1656) and Isaac Casaubon (1559-1614). It does this in order to highlight an oversight in the existing literature concerning Calixtus, and to encourage scholars to revisit the work of Early Modern figures who have previously been considered only from modern disciplinary perspectives. By emphasizing the relationship between Calixtus and Casaubon, this article argues that Calixtus was potentially exposed to much broader circles of intellectual debate than has previously been considered, and that a reevaluation of his work in light of these debates is therefore in order.

CDKN2A mutation in a non-FAMMM kindred with cancers at multiple sites results in a functionally abnormal protein

The CDKN2A gene encodes p16 (CDKN2A), a cell-cycle inhibitor protein which prevents inappropriate cell cycling and, hence, proliferation. Germ-line mutations in CDKN2A predispose to the familial atypical multiple-mole melanoma (FAMMM) syndrome but also have been seen in rare families in which only 1 or 2 individuals are affected by cutaneous malignant melanoma (CMM). We therefore sequenced exons 1alpha and 2 of CDKN2A using lymphocyte DNA isolated from index cases from 67 families with cancers at multiple sites, where the patterns of cancer did not resemble those attributable to known genes such as hMLH1, hMLH2, BRCA1, BRCA2, TP53 or other cancer susceptibility genes. We found one mutation, a mis-sense mutation resulting in a methionine to isoleucine change at codon 53 (M531) of exon 2. The individual tested had developed 2 CMMs but had no dysplastic nevi and lacked a family history of dysplastic nevi or CMM. Other family members had been diagnosed with oral cancer (2 persons), bladder cancer (1 person) and possibly gall-bladder cancer. While this mutation has been reported in Australian and North American melanoma kindreds, we did not observe it in 618 chromosomes from Scottish and Canadian controls. Functional studies revealed that the CDKN2A variant carrying the M531 change was unable to bind effectively to CDK4, showing that this mutation is of pathological significance. Our results have confirmed that CDKN2A mutations are not limited to FAMMM kindreds but also demonstrate that multi-site cancer families without melanoma are very unlikely to contain CDKN2A mutations.

Photoinduced conjugation of Dithioester- and Trithiocarbonate-Functional RAFT polymers with alkenes

We report the photoinduced conjugation of polymers synthesized via reversible addition−fragmentation chain transfer (RAFT) polymerization with a number of low molecular weight (functional) olefins. Upon irradiation of a solution of an aliphatic alkene and the benzyl dithioacetic acid ester (CPDA) or dodecyl trithiocarbonate (DoPAT) functional poly(alkyl acrylate) at the absorption wavelength of the thiocarbonyl group (315 nm), incorporation of the alkene at the polymer chain-end occurred. The most efficient systems identified with regard to the rate of reaction and yield were poly(butyl acrylate)/CPDA/ethyl vinyl ether (78% monoinsertion product after 1 h) and poly(butyl acrylate)/CPDA/1-pentene (73% insertion product after 7 h) at ambient temperature. An in-depth analysis of the reaction mechanism by 1H NMR and online size-exclusion chromatography-electrospray ionization tandem mass spectrometry (SEC/ESI−MSn) revealed that a possible [2 + 2] photoaddition mechanism of conjugation does not take place. Instead, fast β-cleavage of the photoexcited RAFT-end group with subsequent radical addition of an alkene was observed for all employed systems. The presented reaction thus provides a means of spatial and temporal control for the conjugation of alkenes to thiocarbonyl thio-capped macromolecules via the use of UV radiation.

Inferior field loss increases rate of falls in older adults with glaucoma

PURPOSE: To examine the visual predictors of falls and injurious falls among older adults with glaucoma. METHODS: Prospective falls data were collected for 71 community-dwelling adults with primary open-angle glaucoma, mean age 73.9 ± 5.7 years, for one year using monthly falls diaries. Baseline assessment of central visual function included high-contrast visual acuity and Pelli-Robson contrast sensitivity. Binocular integrated visual fields were derived from monocular Humphrey Field Analyser plots. Rate ratios (RR) for falls and injurious falls with 95% confidence intervals (CIs) were based on negative binomial regression models. RESULTS: During the one year follow-up, 31 (44%) participants experienced at least one fall and 22 (31%) experienced falls that resulted in an injury. Greater visual impairment was associated with increased falls rate, independent of age and gender. In a multivariate model, more extensive field loss in the inferior region was associated with higher rate of falls (RR 1.57, 95%CI 1.06, 2.32) and falls with injury (RR 1.80, 95%CI 1.12, 2.98), adjusted for all other vision measures and potential confounding factors. Visual acuity, contrast sensitivity, and superior field loss were not associated with the rate of falls; topical beta-blocker use was also not associated with increased falls risk. CONCLUSIONS: Falls are common among older adults with glaucoma and occur more frequently in those with greater visual impairment, particularly in the inferior field region. This finding highlights the importance of the inferior visual field region in falls risk and assists in identifying older adults with glaucoma at risk of future falls, for whom potential interventions should be targeted. KEY WORDS: glaucoma, visual field, visual impairment, falls, injury

An evaluation of the sustainability of agreements reached through mediations conducted by the dispute resolution centres of the Department of Justice and Attorney General, Queensland between 1999 to 2003

In 1990 the Dispute Resolution Centres Act, 1990 (Qld) (the Act) was passed by the Queensland Parliament. In the second reading speech for the Dispute Resolution Centres Bill on May 1990 the Hon Dean Wells stated that the proposed legislation would make mediation services available “in a non-coercive, voluntary forum where, with the help of trained mediators, the disputants will be assisted towards their own solutions to their disputes, thereby ensuring that the result is acceptable to the parties” (Hansard, 1990, 1718). It was recognised at that time that a method for resolving disputes was necessary for which “the conventional court system is not always equipped to provide lasting resolution” (Hansard, 1990, 1717). In particular, the lasting resolution of “disputes between people in continuing relationships” was seen as made possible through the new legislation; for example, “domestic disputes, disputes between employees, and neighbourhood disputes relating to such issues as overhanging tree branches, dividing fences, barking dogs, smoke, noise and other nuisances are occurring continually in the community” (Hansard, 1990, 1717). The key features of the proposed form of mediation in the Act were articulated as follows: “attendance of both parties at mediation sessions is voluntary; a party may withdraw at any time; mediation sessions will be conducted with as little formality and technicality as possible; the rules of evidence will not apply; any agreement reached is not enforceable in any court; although it could be made so if the parties chose to proceed that way; and the provisions of the Act do not affect any rights or remedies that a party to a dispute has apart from the Act” (Hansard, 1990, 1718). Since the introduction of the Act, the Alternative Dispute Resolution Branch of the Queensland Department of Justice and Attorney General has offered mediation services through, first the Community Justice Program (CJP), and then the Dispute Resolution Centres (DRCs) for a range of family, neighbourhood, workplace and community disputes. These services have mirrored those available through similar government agencies in other states such as the Community Justice Centres of NSW and the Victorian Dispute Resolution Centres. Since 1990, mediation has become one of the fastest growing forms of alternative dispute resolution (ADR). Sourdin has commented that "In addition to the growth in court-based and community-based dispute resolution schemes, ADR has been institutionalised and has grown within Australia and overseas” (2005, 14). In Australia, in particular, the development of ADR service provision “has been assisted by the creation and growth of professional organisations such as the Leading Edge Alternative Dispute Resolvers (LEADR), the Australian Commercial Dispute Centres (ACDC), Australian Disputes Resolution Association (ADRA), Conflict Resolution Network, and the Institute of Arbitrators and Mediators Australia (IAMA)” (Sourdin, 2005, 14). The increased emphasis on the use of ADR within education contexts (particularly secondary and tertiary contexts) has “also led to an increasing acceptance and understanding of (ADR) processes” (Sourdin, 2005, 14). Proponents of the mediation process, in particular, argue that much of its success derives from the inherent flexibility and creativity of the agreements reached through the mediation process and that it is a relatively low cost option in many cases (Menkel-Meadow, 1997, 417). It is also accepted that one of the main reasons for the success of mediation can be attributed to the high level of participation by the parties involved and thus creating a sense of ownership of, and commitment to, the terms of the agreement (Boulle, 2005, 65). These characteristics are associated with some of the core values of mediation, particularly as practised in community-based models as found at the DRCs. These core values include voluntary participation, party self-determination and party empowerment (Boulle, 2005, 65). For this reason mediation is argued as being an effective approach to resolving disputes, that creates a lasting resolution of the issues. Evaluation of the mediation process, particularly in the context of the growth of ADR, has been an important aspect of the development of the process (Sourdin, 2008). Writing in 2005 for example, Boulle, states that “although there is a constant refrain for more research into mediation practice, there has been a not insignificant amount of mediation measurement, both in Australia and overseas” (Boulle, 2005, 575). The positive claims of mediation have been supported to a significant degree by evaluations of the efficiency and effectiveness of the process. A common indicator of the effectiveness of mediation is the settlement rate achieved. High settlement rates for mediated disputes have been found for Australia (Altobelli, 2003) and internationally (Alexander, 2003). Boulle notes that mediation agreement rates claimed by service providers range from 55% to 92% (Boulle, 2005, 590). The annual reports for the Alternative Dispute Resolution Branch of the Queensland Department of Justice and Attorney-General considered prior to the commencement of this study indicated generally achievement of an approximate settlement figure of 86% by the Queensland Dispute Resolution Centres. More recently, the 2008-2009 annual report states that of the 2291 civil dispute mediated in 2007-2008, 86% reached an agreement. Further, of the 2693 civil disputes mediated in 2008-2009, 73% reached an agreement. These results are noted in the report as indicating “the effectiveness of mediation in resolving disputes” and as reflecting “the high level of agreement achieved for voluntary mediations” (Annual Report, 2008-2009, online). Whilst the settlement rates for the DRCs are strong, parties are rarely contacted for long term follow-up to assess whether agreements reached during mediation lasted to the satisfaction of each party. It has certainly been the case that the Dispute Resolution Centres of Queensland have not been resourced to conduct long-term follow-up assessments of mediation agreements. As Wade notes, "it is very difficult to compare "success" rates” and whilst “politicians want the comparison studies (they) usually do not want the delay and expense of accurate studies" (1998, 114). To date, therefore, it is fair to say that the efficiency of the mediation process has been evaluated but not necessarily its effectiveness. Rather, the practice at the Queensland DRCs has been to evaluate the quality of mediation service provision and of the practice of the mediation process. This has occurred, for example, through follow-up surveys of parties' satisfaction rates with the mediation service. In most other respects it is fair to say that the Centres have relied on the high settlement rates of the mediation process as a sign of the effectiveness of mediation (Annual Reports 1991 - 2010). Research of the mediation literature conducted for the purpose of this thesis has also indicated that there is little evaluative literature that provides an in-depth analysis and assessment of the longevity of mediated agreements. Instead evaluative studies of mediation tend to assess how mediation is conducted, or compare mediation with other conflict resolution options, or assess the agreement rate of mediations, including parties' levels of satisfaction with the service provision of the dispute resolution service provider (Boulle, 2005, Chapter 16).

A task-oriented approach to search engine usability studies

Usability is a multi-dimensional characteristic of a computer system. This paper focuses on usability as a measurement of interaction between the user and the system. The research employs a task-oriented approach to evaluate the usability of a meta search engine. This engine encourages and accepts queries of unlimited size expressed in natural language. A variety of conventional metrics developed by academic and industrial research, including ISO standards,, are applied to the information retrieval process consisting of sequential tasks. Tasks range from formulating (long) queries to interpreting and retaining search results. Results of the evaluation and analysis of the operation log indicate that obtaining advanced search engine results can be accomplished simultaneously with enhancing the usability of the interactive process. In conclusion, we discuss implications for interactive information retrieval system design and directions for future usability research. © 2008 Academy Publisher.

Theoretical and numerical investigation of bending properties of Cu nanowires

Based on the AFM-bending experiments, a molecular dynamics (MD) bending simulation model is established which could accurately account for the full spectrum of the mechanical properties of NWs in a double clamped beam configuration, ranging from elasticity to plasticity and failure. It is found that, loading rate exerts significant influence to the mechanical behaviours of nanowires (NWs). Specifically, a loading rate lower than 10 m/s is found reasonable for a homogonous bending deformation. Both loading rate and potential between the tip and the NW are found to play an important role in the adhesive phenomenon. The force versus displacement (F-d) curve from MD simulation is highly consistent in shapes with that from experiments. Symmetrical F-d curves during loading and unloading processes are observed, which reveal the linear-elastic and non-elastic bending deformation of NWs. The typical bending induced tensile-compressive features are observed. Meanwhile, the simulation results are excellently fitted by the classical Euler-Bernoulli beam theory with axial effect. It is concluded that, axial tensile force becomes crucial in bending deformation when the beam size is down to nanoscale for double clamped NWs. In addition, we find shorter NWs will have an earlier yielding and a larger yielding force. Mechanical properties (Young’s modulus & yield strength) obtained from both bending and tensile deformations are found comparable with each other. Specifically, the modulus is essentially similar under these two loading methods, while the yield strength during bending is observed larger than that during tension.

Country of Origin and Internet Publication : Applying the Berne Convention in the Digital Age

Historically, determining the country of origin of a published work presented few challenges, because works were generally published physically – whether in print or otherwise – in a distinct location or few locations. However, publishing opportunities presented by new technologies mean that we now live in a world of simultaneous publication – works that are first published online are published simultaneously to every country in world in which there is Internet connectivity. While this is certainly advantageous for the dissemination and impact of information and creative works, it creates potential complications under the Berne Convention for the Protection of Literary and Artistic Works (“Berne Convention”), an international intellectual property agreement to which most countries in the world now subscribe. Under the Berne Convention’s national treatment provisions, rights accorded to foreign copyright works may not be subject to any formality, such as registration requirements (although member countries are free to impose formalities in relation to domestic copyright works). In Kernel Records Oy v. Timothy Mosley p/k/a Timbaland, et al. however, the Florida Southern District Court of the United States ruled that first publication of a work on the Internet via an Australian website constituted “simultaneous publication all over the world,” and therefore rendered the work a “United States work” under the definition in section 101 of the U.S. Copyright Act, subjecting the work to registration formality under section 411. This ruling is in sharp contrast with an earlier decision delivered by the Delaware District Court in Håkan Moberg v. 33T LLC, et al. which arrived at an opposite conclusion. The conflicting rulings of the U.S. courts reveal the problems posed by new forms of publishing online and demonstrate a compelling need for further harmonization between the Berne Convention, domestic laws and the practical realities of digital publishing. In this article, we argue that even if a work first published online can be considered to be simultaneously published all over the world it does not follow that any country can assert itself as the “country of origin” of the work for the purpose of imposing domestic copyright formalities. More specifically, we argue that the meaning of “United States work” under the U.S. Copyright Act should be interpreted in line with the presumption against extraterritorial application of domestic law to limit its application to only those works with a real and substantial connection to the United States. There are gaps in the Berne Convention’s articulation of “country of origin” which provide scope for judicial interpretation, at a national level, of the most pragmatic way forward in reconciling the goals of the Berne Convention with the practical requirements of domestic law. We believe that the uncertainties arising under the Berne Convention created by new forms of online publishing can be resolved at a national level by the sensible application of principles of statutory interpretation by the courts. While at the international level we may need a clearer consensus on what amounts to “simultaneous publication” in the digital age, state practice may mean that we do not yet need to explore textual changes to the Berne Convention.

A randomized trial of a telephone-delivered exercise intervention for non-urban dwelling women newly diagnosed with breast cancer : exercise for health

Physical activity is important following breast cancer. Trials of non-face-to-face interventions are needed to assist in reaching women living outside major metropolitan areas. This study seeks to evaluate the feasibility and effectiveness of a telephone-delivered, mixed aerobic and resistance exercise intervention for non-urban Australian women with breast cancer. A randomized controlled trial comparing an 8-month intervention delivered by exercise physiologists (n = 73) to usual care (n = 70). Sixty-one percent recruitment rate and 96% retention at 12 months; 79% of women in the intervention group received at least 75% of calls; odds (OR, 95% CI) of meeting intervention targets favored the intervention group for resistance training (OR 3.2; 1.2, 8.9) and aerobic (OR 2.1; 0.8, 5.5) activity. Given the limited availability of physical activity programs for non-urban women with breast cancer, results provide strong support for feasibility and modest support for the efficacy of telephone-delivered interventions.

Safety enhancement of operator protection systems on self-propelled mining equipment

In the long term, with development of skill, knowledge, exposure and confidence within the engineering profession, rigorous analysis techniques have the potential to become a reliable and far more comprehensive method for design and verification of the structural adequacy of OPS, write Nimal J Perera, David P Thambiratnam and Brian Clark. This paper explores the potential to enhance operator safety of self-propelled mechanical plant subjected to roll over and impact of falling objects using the non-linear and dynamic response simulation capabilities of analytical processes to supplement quasi-static testing methods prescribed in International and Australian Codes of Practice for bolt on Operator Protection Systems (OPS) that are post fitted. The paper is based on research work carried out by the authors at the Queensland University of Technology (QUT) over a period of three years by instrumentation of prototype tests, scale model tests in the laboratory and rigorous analysis using validated Finite Element (FE) Models. The FE codes used were ABAQUS for implicit analysis and LSDYNA for explicit analysis. The rigorous analysis and dynamic simulation technique described in the paper can be used to investigate the structural response due to accident scenarios such as multiple roll over, impact of multiple objects and combinations of such events and thereby enhance the safety and performance of Roll Over and Falling Object Protection Systems (ROPS and FOPS). The analytical techniques are based on sound engineering principles and well established practice for investigation of dynamic impact on all self propelled vehicles. They are used for many other similar applications where experimental techniques are not feasible.

Developing tools to facilitate integrated reflection

Much has been said and documented about the key role that reflection can play in the ongoing development of e-portfolios, particularly e-portfolios utilised for teaching and learning. A review of e-portfolio platforms reveals that a designated space for documenting and collating personal reflections is a typical design feature of both open source and commercial off-the-shelf software. Further investigation of tools within e-portfolio systems for facilitating reflection reveals that, apart from enabling personal journalism through blogs or other writing, scaffolding tools that encourage the actual process of reflection are under-developed. Investigation of a number of prominent e-portfolio projects also reveals that reflection, while presented as critically important, is often viewed as an activity that takes place after a learning activity or experience and not intrinsic to it. This paper assumes an alternative, richer conception of reflection: a process integral to a wide range of activities associated with learning, such as inquiry, communication, editing, analysis and evaluation. Such a conception is consistent with the literature associated with ‘communities of practice’, which is replete with insight into ‘learning through doing’, and with a ‘whole minded’ approach to inquiry. Thus, graduates who are ‘reflective practitioners’ who integrate reflection into their learning will have more to offer a prospective employer than graduates who have adopted an episodic approach to reflection. So, what kinds of tools might facilitate integrated reflection? This paper outlines a number of possibilities for consideration and development. Such tools do not have to be embedded within e-portfolio systems, although there are benefits in doing so. In order to inform future design of e-portfolio systems this paper presents a faceted model of knowledge creation that depicts an ‘ecology of knowing’ in which interaction with, and the production of, learning content is deepened through the construction of well-formed questions of that content. In particular, questions that are initiated by ‘why’ are explored because they are distinguished from the other ‘journalist’ questions (who, what, when, where, and where) in that answers to them demand explanative, as opposed to descriptive, content. They require a rationale. Although why questions do not belong to any one genre and are not simple to classify — responses can contain motivational, conditional, causal, and/or existential content — they do make a difference in the acquisition of understanding. The development of scaffolding that builds on why-questioning to enrich learning is the motivation behind the research that has informed this paper.