Development of a high-level gene expression system for the production of bioplastics in sugarcane

Despite various approaches, the production of biodegradable plastics such as polyhydroxybutyrate (PHB) in transgenic plants has met with limited success due largely to low expression levels. Even in the few instances where high levels of protein expression have been reported, the transgenic plants have been stunted indicating PHB is phytotoxic (Poirier 2002). This PhD describes the application of a novel virus-based gene expression technology, termed InPAct („In Plant Activation.), for the production of PHB in tobacco and sugarcane. InPAct is based on the rolling circle replication mechanism by which circular ssDNA viruses replicate and provides a system for controlled, high-level gene expression. Based on these features, InPAct was thought to represent an ideal system to enable the controlled, high-level expression of the three phb genes (phbA, phbB and phbC) required for PHB production in sugarcane at a preferred stage of plant growth. A Tobacco yellow dwarf virus (TbYDV)-based InPAct-phbA vector, as well as linear vectors constitutively expressing phbB and phbC were constructed and different combinations were used to transform tobacco leaf discs. A total of four, eight, three and three phenotypically normal tobacco lines were generated from discs transformed with InPAct-phbA, InPAct-phbA + p1300-TaBV P-phbB/phbC- 35S T, p1300-35S P-phbA-NOS T + p1300-TaBV P-phbB/phbC-35S T and InPAct-GUS, respectively. To determine whether the InPAct cassette could be activated in the presence of the TbYDV Rep, leaf samples from the eight InPActphbA + p1300-TaBV P-phbB/phbC-35S T plants were agroinfiltrated with p1300- TbYDV-Rep/RepA. Three days later, successful activation was indicated by the detection of episomes using both PCR and Southern analysis. Leaf discs from the eight InPAct-phbA + p1300-TaBV P-phbB/phbC-35S T transgenic plant lines were agroinfiltrated with p1300-TbYDV-Rep/RepA and leaf tissue was collected ten days post-infiltration and examined for the presence of PHB granules. Confocal microscopy and TEM revealed the presence of typical PHB granules in five of the eight lines, thus demonstrating the functionality of InPActbased PHB production in tobacco. However, analysis of leaf extracts by HPLC failed to detect the presence of PHB suggesting only very low level expression levels. Subsequent molecular analysis of three lines revealed low levels of correctly processed mRNA from the catalase intron contained within the InPAct cassette and also the presence of cryptic splice sites within the intron. In an attempt to increase expression levels, new InPAct-phb cassettes were generated in which the castorbean catalase intron was replaced with a synthetic intron (syntron). Further, in an attempt to both increase and better control Rep/RepA-mediated activation of InPAct cassettes, Rep/RepA expression was placed under the control of a stably integrated alc switch. Leaf discs from a transgenic tobacco line (Alc ML) containing 35S P-AlcR-AlcA P-Rep/RepA were supertransformed with InPAct-phbAsyn or InPAct-GUSsyn using Agrobacterium and three plants (lines) were regenerated for each construct. Analysis of the RNA processing of the InPAct-phbAsyn cassette revealed highly efficient and correct splicing of the syntron, thus supporting its inclusion within the InPAct system. To determine the efficiency of the alc switch to activate InPAct, leaf material from the three Alc ML + InPAct-phbAsyn lines was either agroinfiltrated with 35S P-Rep/RepA or treated with ethanol. Unexpectedly, episomes were detected not only in the infiltrated and ethanol treated samples, but also in non-treated samples. Subsequent analysis of transgenic Alc ML + InPAct-GUS lines, confirmed that the alc switch was leaky in tissue culture. Although this was shown to be reversible once plants were removed from the tissue culture environment, it made the regeneration of Alc ML + InPAct-phbsyn plant lines extremely difficult, due to unintentional Rep expression and therefore high levels of phb expression and phytotoxic PHB production. Two Alc ML + InPAct-phbAsyn + p1300-TaBV P-phbB/phbC-35S T transgenic lines were able to be regenerated, and these were acclimatised, alcohol-treated and analysed. Although episome formation was detected as late as 21 days post activation, no PHB was detected in the leaves of any plants using either microscopy or HPLC, suggesting the presence of a corrupt InPAct-phbA cassette in both lines. The final component of this thesis involved the application of both the alc switch and the InPAct systems to sugarcane in an attempt to produce PHB. Initial experiments using transgenic Alc ML + InPAct-GUS lines indicated that the alc system was not functional in sugarcane under the conditions tested. The functionality of the InPAct system, independent of the alc gene switch, was subsequently examined by bombarding the 35S Rep/RepA cassette into leaf and immature leaf whorl cells derived from InPAct-GUS transgenic sugarcane plants. No GUS expression was observed in leaf tissue, whereas weak and irregular GUS expression was observed in immature leaf whorl tissue derived from two InPAct- GUS lines and two InPAct-GUS + 35S P-AlcR-AlcA P-GUS lines. The most plausible reason to explain the inconsistent and low levels of GUS expression in leaf whorls is a combination of low numbers of sugarcane cells in the DNA replication-conducive S-phase and the irregular and random nature of sugarcane cells bombarded with Rep/RepA. This study details the first report to develop a TbYDV-based InPAct system under control of the alc switch to produce PHB in tobacco and sugarcane. Despite the inability to detect quantifiable levels of PHB levels in either tobacco or sugarcane, the findings of this study should nevertheless assist in the further development of both the InPAct system and the alc system, particularly for sugarcane and ultimately lead to an ethanol-inducible InPAct gene expression system for the production of bioplastics and other proteins of commercial value in plants.

Nanostructured mesoporous bioglass coated zirconia scaffolds for bone tissue engineering

For the filling and reconstruction of non-healing bone defects, the application of porous ceramic scaffold as bone substitutes is considered to be a reasonable choice. In bone tissue engineering, an ideal scaffold must satisfy several criterias such as open porosity, having high compressive strength (it depends where in body, and if external fixatures are used) and the practicability for cell migration. Many researchers have focused on enhancing the mechanical properties of hydroxyapatite scaffolds by combining it with other biomaterials, such as bioglass and polymers. Nevertheless, there is still a lack of suitable scaffolds based on porous biomaterials. In this study, zirconia scaffolds from two different templates (polyurethane (PU) and Acrylonitrile Butadiene Styrene (ABS) templates) were successfully fabricated with dissimilar fabrication techniques. The scaffold surfaces were further modified with mesoporous bioglass for the purpose of bone tissue engineering. In the study of PU template scaffold, high porosity (~88%) sol-gel derived yttria-stabilized zirconia (YSZ) scaffold was prepared by a polyurethane (PU) foam replica method using sol-gel derived zirconia for the first time, and double coated with Mesoporous Bioglass (MBGs) coating. For the ABS template scaffold, two types of templates (cube and cylinder) with different strut spacings were used and fabricated by a 3D Rapid Prototyper. Subsequently, zirconia scaffolds with low porosity (63±2.8% to 68±2.5%) were fabricated by embedding the zirconia powder slurry into the ABS templates and burning out the ABS to produce a uniform porous structure. The zirconia scaffolds were double coated with mesoporous bioglass by dip coating for the first time. The porosities of the scaffolds were calculated before and after coating. The microstructures were then examined using scanning electron microscopy and the mechanical properties were evaluated using compressive test. Accordingly, relationships between microstructure, processing and mechanical behaviour of the porous zirconia was discussed. Scaffold biocompatibility and bioactivity was also evaluated using a bone marrow stromal cell (BMSC) proliferation test and a simulated body fluid test.

Investigation of methods for increasing the energy efficiency on unmanned aerial vehicles (UAVS)

In recent years, development of Unmanned Aerial Vehicles (UAV) has become a significant growing segment of the global aviation industry. These vehicles are developed with the intention of operating in regions where the presence of onboard human pilots is either too risky or unnecessary. Their popularity with both the military and civilian sectors have seen the use of UAVs in a diverse range of applications, from reconnaissance and surveillance tasks for the military, to civilian uses such as aid relief and monitoring tasks. Efficient energy utilisation on an UAV is essential to its functioning, often to achieve the operational goals of range, endurance and other specific mission requirements. Due to the limitations of the space available and the mass budget on the UAV, it is often a delicate balance between the onboard energy available (i.e. fuel) and achieving the operational goals. This thesis presents an investigation of methods for increasing the energy efficiency on UAVs. One method is via the development of a Mission Waypoint Optimisation (MWO) procedure for a small fixed-wing UAV, focusing on improving the onboard fuel economy. MWO deals with a pre-specified set of waypoints by modifying the given waypoints within certain limits to achieve its optimisation objectives of minimising/maximising specific parameters. A simulation model of a UAV was developed in the MATLAB Simulink environment, utilising the AeroSim Blockset and the in-built Aerosonde UAV block and its parameters. This simulation model was separately integrated with a multi-objective Evolutionary Algorithm (MOEA) optimiser and a Sequential Quadratic Programming (SQP) solver to perform single-objective and multi-objective optimisation procedures of a set of real-world waypoints in order to minimise the onboard fuel consumption. The results of both procedures show potential in reducing fuel consumption on a UAV in a ight mission. Additionally, a parallel Hybrid-Electric Propulsion System (HEPS) on a small fixedwing UAV incorporating an Ideal Operating Line (IOL) control strategy was developed. An IOL analysis of an Aerosonde engine was performed, and the most efficient (i.e. provides greatest torque output at the least fuel consumption) points of operation for this engine was determined. Simulation models of the components in a HEPS were designed and constructed in the MATLAB Simulink environment. It was demonstrated through simulation that an UAV with the current HEPS configuration was capable of achieving a fuel saving of 6.5%, compared to the ICE-only configuration. These components form the basis for the development of a complete simulation model of a Hybrid-Electric UAV (HEUAV).

Combining influence maps and cellular automata for reactive game agents

Agents make up an important part of game worlds, ranging from the characters and monsters that live in the world to the armies that the player controls. Despite their importance, agents in current games rarely display an awareness of their environment or react appropriately, which severely detracts from the believability of the game. Some games have included agents with a basic awareness of other agents, but they are still unaware of important game events or environmental conditions. This paper presents an agent design we have developed, which combines cellular automata for environmental modeling with influence maps for agent decision-making. The agents were implemented into a 3D game environment we have developed, the EmerGEnT system, and tuned through three experiments. The result is simple, flexible game agents that are able to respond to natural phenomena (e.g. rain or fire), while pursuing a goal.

Scripting versus emergence : issues for game developers and players in game environment design

This paper defines and discusses two contrasting approaches to designing game environments. The first, referred to as scripting, requires developers to anticipate, hand-craft and script specific game objects, events and player interactions. The second, known as emergence, involves defining general, global rules that interact to give rise to emergent gameplay. Each of these approaches is defined, discussed and analyzed with respect to the considerations and affects for game developers and game players. Subsequently, various techniques for implementing these design approaches are identified and discussed. It is concluded that scripting and emergence are two extremes of the same continuum, neither of which are ideal for game development. Rather, there needs to be a compromise in which the boundaries of action (such as story and game objectives) can be hardcoded and non-scripted behavior (such as interactions and strategies) are able to emerge within these boundaries.

Guidance of aircraft in periodic inspection tasks

This paper presents a guidance approach for aircraft in periodic inspection tasks. The periodic inspection task involves flying to a series of desired fixed points of inspection with specified attitude requirements so that requirements for downward looking sensors, such as cameras, are achieved. We present a solution using a precision guidance law and a bank turn dynamics model. High fidelity simulation studies illustrate the effectiveness of this approach under both ideal (nil-wind) and non-ideal (wind) conditions.

Using mobile phones to improve offline access to online information : distributed content delivery

Mobile phones are now powerful and pervasive making them ideal information browsers. The Internet has revolutionized our lives and is a major knowledge sharing media. However, many mobile phone users cannot access the Internet (for financial or technical reasons) and so the mobile Internet has not been fully realized. We propose a novel content delivery network based on both a factual and speculative analysis of today’s technology and analyze its feasibility. If adopted people living in remote regions without Internet will be able to access essential (static) information with periodic updates.

How design of the physical environment impacts early learning : educators and parents perspectives

This research explores the quality and importance of the physical environment of two early learning centres on the Sunshine Coast in Queensland, utilising qualitative interviews with parents (n=4) and educators (n=4) to understand how design might impact on children’s development and a quantitative rating (the Early Childhood Physical Environment Rating Scale; ECPERS) to assess the quality of the physical built environment and infrastructure. With an average ECPERS quality rating, thematic analysis of the interviews revealed that educators and parents viewed the physical environment as important to a child’s development, although the quality of staff was predominant. Early learning centres should be ‘homely’, inviting, bright and linked to the outdoors, with participants describing how space “welcomes the child, makes them feel safe and encourages learning”. Four key themes characterised views: Emotional Connection (quality of staff and physical environment), Experiencing Design (impact of design on child development), Hub for Community Integration (relationships and resources) and Future Vision (ideal physical environment, technology and ratings). With participants often struggling to clearly articulate their thoughts on design issues, a collaborative and jargon-free approach to designing space is required. These findings will help facilitate discussion about the role and design of the physical environment in early childhood centres, with the tangible examples of ‘ideal space’ enhancing communication between architects and educators about how best to design and reconfigure space to enhance learning outcomes.

Breastfeeding self-efficacy and alternative techniques to overcome maternal or infant breastfeeding challenges : a retrospective descriptive study

Background: Breastfeeding is the internationally accepted ideal in infant feeding. Ensuring mothers and babies receive optimal benefits, in both the short and long term, is dependent upon the successful establishment of breastfeeding in the first week. Many maternal and infant challenges can occur during the establishment of breastfeeding (Lactogenesis II). There are also many methods and devices (alternative techniques) which can be used to help, but the majority do not have an evidence-base. The mother.s self-confidence (self-efficacy) can be challenged by these unexpected circumstances, but understanding of the relationship is unclear. Method: This descriptive study used mail survey (including the Breastfeeding Self-Efficacy Scale . Short Form) to obtain the mother.s reports of their self-efficacy and their breastfeeding experience during the first week following birth, as well as actual use of alternative techniques. This study included all mothers of full term healthy singleton infants from one private hospital in Brisbane who began any breastfeeding. The data collection took place from November 2008 to February 2009. Ethical approval was granted from the research site and QUT Human Research Ethics Committee. Results: A total of 128 questionnaires were returned, a response rate of 56.9%. The sample was dissimilar to the Queensland population with regard to age, income, and education level, all of which were higher in this study. The sample was similar to the Queensland population in terms of parity and marital status. The rate of use of alternative techniques was 48.3%. The mean breastfeeding self-efficacy score of those who used any alternative technique was 43.43 (SD=12.19), and for those who did not, it was 58.32 (SD=7.40). Kruskal-Wallis analysis identified that the median self efficacy score for those who used alternative techniques was significantly lower than median self efficacy scores for those who did not use alternative techniques. The reasons women used alternative techniques varied widely, and their knowledge of alternative techniques was good. Conclusion: This study is the first to document breastfeeding self-efficacy of women who used alternative techniques to support their breastfeeding goals in the first week postpartum. An individualised clinical intervention to develop women.s self-efficacy with breastfeeding is important to assist mother/infant dyads encountering challenges to breastfeeding in the first week postpartum.

Is the public sector ready to collaborate? Human resource implications of collaborative Arrangements

Relational governance arrangements across agencies and sectors have become prevalent as a means for government to become more responsive and effective in addressing complex, large scale or ‘wicked’ problems. The primary characteristic of such ‘collaborative’ arrangements is the utilisation of the joint capacities of multiple organisations to achieve collaborative advantage, which Huxham (1993) defines as the attainment of creative outcomes that are beyond the ability of single agencies to achieve. Attaining collaborative advantage requires organisations to develop collaborative capabilities that prepare organisations for collaborative practice (Huxham, 1993b). Further, collaborations require considerable investment of staff effort that could potentially be used beneficially elsewhere by both the government and non-government organisations involved in collaboration (Keast and Mandell, 2010). Collaborative arrangements to deliver services therefore requires a reconsideration of the way in which resources, including human resources, are conceptualised and deployed as well as changes to both the structure of public service agencies and the systems and processes by which they operate (Keast, forthcoming). A main aim of academic research and theorising has been to explore and define the requisite characteristics to achieve collaborative advantage. Such research has tended to focus on definitional, structural (Turrini, Cristofoli, Frosini, & Nasi, 2009) and organisational (Huxham, 1993) aspects and less on the roles government plays within cross-organisational or cross-sectoral arrangements. Ferlie and Steane (2002) note that there has been a general trend towards management led reforms of public agencies including the HRM practices utilised. Such trends have been significantly influenced by New Public Management (NPM) ideology with limited consideration to the implications for HRM practice in collaborative, rather than market contexts. Utilising case study data of a suite of collaborative efforts in Queensland, Australia, collected over a decade, this paper presents an examination of the network roles government agencies undertake. Implications for HRM in public sector agencies working within networked arrangements are drawn and implications for job design, recruitment, deployment and staff development are presented. The paper also makes theoretical advances in our understanding of Strategic Human Resource Management (SHRM) in network settings. While networks form part of the strategic armoury of government, networks operate to achieve collaborative advantage. SHRM with its focus on competitive advantage is argued to be appropriate in market situations, however is not an ideal conceptualisation in network situations. Commencing with an overview of literature on networks and network effectiveness, the paper presents the case studies and methodology; provides findings from the case studies in regard to the roles of government to achieve collaborative advantage and implications for HRM practice are presented. Implications for SHRM are considered.

Learning by design workshop

The Learning by Design Workshop Program 2010, a part of the Queensland Government Unlimited: Designing for the Asia Pacific Event Program, was a one-day professional development design thinking workshop run on October 9, 2011 at The Edge, State Library of Queensland for self-selected public and private secondary school teachers from the subject areas of Visual Art, Graphics and Industrial Technology and Design. Participants were drawn from a database of Brisbane and regional Queensland schools from the goDesign and Living City Workshop Programs. It aimed to generate leadership within schools for design-led education and creative thinking and give teachers a rare opportunity to work with professional designers to generate future strategies for design-based learning. Teachers were introduced to the concept of design thinking in education by international keynote speakers CJ Lim (Studio 8 Architects) and Jeb Brugmann (The Next Practice), national speaker Oliver Freeman (NevilleFreeman Agency) and three Queensland speakers, Alexander Loterztain, David Williams and Keith Holledge. Inspired by the Unlimited showcase exhibition Make Change: Design Thinking in Action and ‘Idea Starters’/teaching resources provided, teachers worked with a professional designer (from a discipline of architecture, interior design, industrial design, urban design, graphic design or landscape architecture) in ten random teams, to generate optimistic ideas for the Ideal City of tomorrow, each considering a theme – Food, Water, Transport, Ageing, Growth, Employment, Shelter, Health, Education and Energy. They then discussed how this process could be best activated and expanded on to build interest and knowledge in design thinking in the classroom. Assisted by illustrators, the teams prepared a visual presentation of their ideas and process from art materials provided. The workshop culminated in a video-taped interactive design charette to the larger group, which is intended to be utilised as a toolkit and praxis for teachers as part of the State Library of Queensland Design Minds Website Project.

Generation workshop 2010

The Generation Workshop Program 2010, a part of the Queensland Government Unlimited: Designing for the Asia Pacific Event Program, consisted of two one-day intensive design thinking workshops run on October 7-8, 2011 at The Edge, State Library of Queensland, for 100 senior secondary students and 20 secondary teachers self-selected from the subject areas of Visual Art, Graphics and Industrial Technology and Design. Participants were drawn from a database of Brisbane and regional Queensland private and public schools from the goDesign and Living City Workshop Programs. The workshop aimed to facilitate awareness in young people of the role of design in society and the value of design thinking skills in solving complex problems facing the Asia Pacific Region, and to inspire the generation of strategies for our future cities. It also aimed to encourage the collaboration of professional designers with secondary schools to inspire post-secondary pathways and idea generation for education. Inspired by international and national speakers Bunker Roy (Barefoot College) and Hael Kobayashi (Associate Producer on "Happy Feet" film for Australia's Animal Logic), the Unlimited showcase exhibition Make Change: Design Thinking in Action and ‘Idea Starters’/teaching resources provided, students worked with a teacher in ten random teams, to generate optimistic strategies for the Ideal City of tomorrow, each considering a theme – Food, Water, Transport, Ageing, Growth, Employment, Shelter, Health, Education and Energy. Each team of 6 was led by a professional designer (from the discipline of architecture, interior design, industrial design, urban design, graphic design or landscape architecture) who was a catalyst for driving the student creative thinking process. Assisted by illustrators, the teams prepared a visual presentation of their idea from art materials provided. The workshop culminated in a video-taped interactive design chatter to the larger group, which will be utilised as a toolkit and praxis for teachers as part of the State Library of Queensland Design Minds Project. Photos of student design work were published on the Unlimited website.

A recommendation method for online dating networks based on social relations and demographic information

A new relationship type of social networks - online dating - are gaining popularity. With a large member base, users of a dating network are overloaded with choices about their ideal partners. Recommendation methods can be utilized to overcome this problem. However, traditional recommendation methods do not work effectively for online dating networks where the dataset is sparse and large, and a two-way matching is required. This paper applies social networking concepts to solve the problem of developing a recommendation method for online dating networks. We propose a method by using clustering, SimRank and adapted SimRank algorithms to recommend matching candidates. Empirical results show that the proposed method can achieve nearly double the performance of the traditional collaborative filtering and common neighbor methods of recommendation.

Closing the loop 21st century style : providing feedback on written assessment via MP3 recordings

Feedback on student performance, whether in the classroom or on written assignments, enables them to reflect on their understandings and restructure their thinking in order to develop more powerful ideas and capabilities. Research has identified a number of broad principles of good feedback practice. These include the provision of feedback that facilitates the development of reflection in learning; helps clarify what good performance is in terms of goals, criteria and expected standards; provides opportunities to close the gap between current and desired performance; delivers high quality information to students about their learning; and encourages positive motivational beliefs and self-esteem. However, high staff–student ratios and time pressures often result in a gulf between this ideal and reality. Whilst greater use of criteria referenced assessment has enabled an improvement in the extent of feedback being provided to students, this measure alone does not go far enough to satisfy the requirements of good feedback practice. Technology offers an effective and efficient means by which personalised feedback may be provided to students. This paper presents the findings of a trial of the use of the freely available Audacity program to provide individual feedback via MP3 recordings to final year Media Law students at the Queensland University of Technology on their written assignments. The trial has yielded wide acclaim by students as an effective means of explaining the exact reasons why they received the marks they were awarded, the things they did well and the areas needing improvement. It also showed that good feedback practice can be achieved without the burden of an increase in staff workload.

Comparison of three-dimensional profiles over time

In this paper, we describe an analysis for data collected on a three-dimensional spatial lattice with treatments applied at the horizontal lattice points. Spatial correlation is accounted for using a conditional autoregressive model. Observations are defined as neighbours only if they are at the same depth. This allows the corresponding variance components to vary by depth. We use the Markov chain Monte Carlo method with block updating, together with Krylov subspace methods, for efficient estimation of the model. The method is applicable to both regular and irregular horizontal lattices and hence to data collected at any set of horizontal sites for a set of depths or heights, for example, water column or soil profile data. The model for the three-dimensional data is applied to agricultural trial data for five separate days taken roughly six months apart in order to determine possible relationships over time. The purpose of the trial is to determine a form of cropping that leads to less moist soils in the root zone and beyond.We estimate moisture for each date, depth and treatment accounting for spatial correlation and determine relationships of these and other parameters over time.