Investigation of highly regulated promoters for the production of recombinant proteins in plants

Plants have been identified as promising expression systems for the commercial production of recombinant proteins. Plant-based protein production or “biofarming” offers a number of advantages over traditional expression systems in terms of scale of production, the capacity for post-translation processing, providing a product free of contaminants and cost effectiveness. A number of pharmaceutically important and commercially valuable proteins, such as antibodies, biopharmaceuticals and industrial enzymes are currently being produced in plant expression systems. However, several challenges still remain to improve recombinant protein yield with no ill effect on the host plant. The ability for transgenic plants to produce foreign proteins at commercially viable levels can be directly related to the level and cell specificity of the selected promoter driving the transgene. The accumulation of recombinant proteins may be controlled by a tissue-specific, developmentally-regulated or chemically-inducible promoter such that expression of recombinant proteins can be spatially- or temporally- controlled. The strict control of gene expression is particularly useful for proteins that are considered toxic and whose expression is likely to have a detrimental effect on plant growth. To date, the most commonly used promoter in plant biotechnology is the cauliflower mosaic virus (CaMV) 35S promoter which is used to drive strong, constitutive transgene expression in most organs of transgenic plants. Of particular interest to researchers in the Centre for Tropical Crops and Biocommodities at QUT are tissue-specific promoters for the accumulation of foreign proteins in the roots, seeds and fruit of various plant species, including tobacco, banana and sugarcane. Therefore this Masters project aimed to isolate and characterise root- and seed-specific promoters for the control of genes encoding recombinant proteins in plant-based expression systems. Additionally, the effects of matching cognate terminators with their respective gene promoters were assessed. The Arabidopsis root promoters ARSK1 and EIR1 were selected from the literature based on their reported limited root expression profiles. Both promoters were analysed using the PlantCARE database to identify putative motifs or cis-acting elements that may be associated with this activity. A number of motifs were identified in the ARSK1 promoter region including, WUN (wound-inducible), MBS (MYB binding site), Skn-1, and a RY core element (seed-specific) and in the EIR1 promoter region including, Skn-1 (seed-specific), Box-W1 (fungal elicitor), Aux-RR core (auxin response) and ABRE (ABA response). However, no previously reported root-specific cis-acting elements were observed in either promoter region. To confirm root specificity, both promoters, and truncated versions, were fused to the GUS reporter gene and the expression cassette introduced into Arabidopsis via Agrobacterium-mediated transformation. Despite the reported tissue-specific nature of these promoters, both upstream regulatory regions directed constitutive GUS expression in all transgenic plants. Further, similar levels of GUS expression from the ARSK1 promoter were directed by the control CaMV 35S promoter. The truncated version of the EIR1 promoter (1.2 Kb) showed some differences in the level of GUS expression compared to the 2.2 Kb promoter. Therefore, this suggests an enhancer element is contained in the 2.2 Kb upstream region that increases transgene expression. The Arabidopsis seed-specific genes ATS1 and ATS3 were selected from the literature based on their seed-specific expression profiles and gene expression confirmed in this study as seed-specific by RT-PCR analysis. The selected promoter regions were analysed using the PlantCARE database in order to identify any putative cis elements. The seed-specific motifs GCN4 and Skn-1 were identified in both promoter regions that are associated with elevated expression levels in the endosperm. Additionaly, the seed-specific RY element and the ABRE were located in the ATS1 promoter. Both promoters were fused to the GUS reporter gene and used to transform Arabidopsis plants. GUS expression from the putative promoters was consitutive in all transgenic Arabidopsis tissue tested. Importantly, the positive control FAE1 seed-specific promoter also directed constitutive GUS expression throughout transgenic Arabidopsis plants. The constitutive nature seen in all of the promoters used in this study was not anticipated. While variations in promoter activity can be caused by a number of influencing factors, the variation in promoter activity observed here would imply a major contributing factor common to all plant expression cassettes tested. All promoter constructs generated in this study were based on the binary vector pCAMBIA2300. This vector contains the plant selection gene (NPTII) under the transcriptional control of the duplicated CaMV 35S promoter. This CaMV 35S promoter contains two enhancer domains that confer strong, constitutive expression of the selection gene and is located immediately upstream of the promoter-GUS fusion. During the course of this project, Yoo et al. (2005) reported that transgene expression is significantly affected when the expression cassette is located on the same T-DNA as the 35S enhancer. It was concluded, the trans-acting effects of the enhancer activate and control transgene expression causing irregular expression patterns. This phenomenon seems the most plausible reason for the constitutive expression profiles observed with the root- and seed-specific promoters assessed in this study. The expression from some promoters can be influenced by their cognate terminator sequences. Therefore, the Arabidopsis ARSK1, EIR1, ATS1 and ATS3 terminator sequences were isolated and incorporated into expression cassettes containing the GUS reporter gene under the control of their cognate promoters. Again, unrestricted GUS activity was displayed throughout transgenic plants transformed with these reporter gene fusions. As previously discussed constitutive GUS expression was most likely due to the trans-acting effect of the upstream CaMV 35S promoter in the selection cassette located on the same T-DNA. The results obtained in this study make it impossible to assess the influence matching terminators with their cognate promoters have on transgene expression profiles. The obvious future direction of research continuing from this study would be to transform pBIN-based promoter-GUS fusions (ie. constructs containing no CaMV 35S promoter driving the plant selection gene) into Arabidopsis in order to determine the true tissue specificity of these promoters and evaluate the effects of their cognate 3’ terminator sequences. Further, promoter truncations based around the cis-elements identified here may assist in determining whether these motifs are in fact involved in the overall activity of the promoter.

Studio game-on

The studio-gameon event was supported by the Institute of the Creative Industries and Innovation and the Faculty of IT as part of the State Library of Queensland GAME ON exhibition (ex Barbican, UK) The studio produced a full game in six weeks. It was a curated event, a live web-based exhibition, a performance for the public and the team produced a digital / creative work which is available for download. The studio enabled a team of students to experience the pressures of a real game studio within the space of the precincts but also very much in the public eye. It was a physical hypothesis of the University's mantra - "for the real world" statement: Studio GameOn is an opportunity running alongside the GAME ON exhibition at the State Library of Queensland. The exhibition itself is open to the public from November 17th through to February 15th. The studio runs from January 5th to February 13th 2009. The Studio GameOn challenge? To put together a team of game developers and make a playable game in six weeks! The studio-game on team consists of a group of game developers in training - the team members are all students who are either half-way through or completing a qualification in game design and all its elements - we have designers, artists, programmers and productionteam members. We are also fortunate to have an Industry Board consisting of local Queensland Games professionals: John Passfield (Red Sprite Studios), Adrian Cook (WIldfire Studios) and Duncan Curtis and Marko Grgic (The 3 Blokes). We also invite the public to play with us - there is an ideas box both on-site at the State Library and a number of ways to communicate with us on this studio website.

Effects of age on peripheral ocular aberrations

On-axis monochromatic higher-order aberrations increase with age. Few studies have been made of peripheral refraction along the horizontal meridian of older eyes, and none of their off-axis higher-order aberrations. We measured wave aberrations over the central 42°x32° visual field for a 5mm pupil in 10 young and 7 older emmetropes. Patterns of peripheral refraction were similar in the two groups. Coma increased linearly with field angle at a significantly higher rate in older than in young emmetropes (−0.018±0.007 versus −0.006±0.002 µm/deg). Spherical aberration was almost constant over the measured field in both age groups and mean values across the field were significantly higher in older than in young emmetropes (+0.08±0.05 versus +0.02±0.04 µm). Total root-mean-square and higher-order aberrations increased more rapidly with field angle in the older emmetropes. However, the limits to monochromatic peripheral retinal image quality are largely determined by the second-order aberrations, which do not change markedly with age, and under normal conditions the relative importance of the increased higher-order aberrations in older eyes is lessened by the reduction in pupil diameter with age. Therefore it is unlikely that peripheral visual performance deficits observed in normal older individuals are primarily attributable to the increased impact of higher-order aberration.

Venture creation and resource processes : using Bricolage sustainability ventures

Principal Topic: Resource decisions are critical to the venture creation process, which has important subsequent impacts on venture creation and performance (Boeker, 1989). Most entrepreneurs however, suffer substantial resource constraints in venture creation and during venture growth (Shepherd et al., 2000). Little is known about how high potential, sustainability ventures (the ventures of interest in this research), despite resource constraints, achieve continued venture persistence and venture success. One promising theory that explicitly links to resource constraints is a concept developed by Levi Strauss (1967) termed bricolage. Bricolage aligns with notions of resourcefulness: using what's on hand, through making do, and recombining resources for new or novel purposes (Baker & Nelson 2005). To the best of our knowledge, previous studies have not systematically investigated internal and external constraints, their combinations, and subsequent bricolage patterns. The majority of bricolage literature focuses on external environmental constraints (e.g. Wieck 1989; Baker & Nelson 2005), thereby paying less attention to in evaluating internal constraints (e.g. skills and capabilities) or constraint combinations. In this paper we focus on ventures that typically face resource-poor environments. High potential, nascent and young sustainability ventures are often created and developed with resource constraints and in some cases, have greater resource requirements owing to higher levels of technical sophistication of their products (Rothaermel & Deeds 2006). These ventures usually have high aspirations and potential for growth who ''seeks to meet the needs and aspirations without compromising the ability to meet those of the future'' (Brundtland Commission 1983). High potential ventures are increasingly attributed with a central role in the development of innovation, and employment in developed economies (Acs 2008). Further, increasing awareness of environmental and sustainability issues has fostered demand for business processes that reduce detrimental environmental impacts of global development (Dean & McMullen 2007) and more environmentally sensitive products and services: representing an opportunity for the development of ventures that seek to satisfy this demand through entrepreneurial action. These ventures may choose to ''make do'' with existing resources in developing resource combinations that produce the least impact on the environment. The continuous conflict between the greater requirements for resources and limited resource availability in high potential sustainable ventures, with the added complexity of balancing this with an uncompromising focus on using ''what's on hand'' to lessen environment impacts may make bricolage behaviours critical for these ventures. Research into bricolage behaviour is however, the exception rather than the rule (Cunha 2005). More research is therefore needed to further develop and extend this emerging concept, especially in the context of sustainability ventures who are committed to personal and social goals of resourcefulness. To date, however, bricolage has not been studied specifically among high potential sustainable ventures. This research seeks to develop an in depth understanding of the impact of internal and external constraints and their combinations on the mechanisms employed in bricolage behaviours in differing dynamic environments. The following research question was developed to investigate this: How do internal, external resource constraints (or their combinations) impact bricolage resource decisions in high potential sustainability ventures? ---------- Methodology/Key Propositions: 6 case studies will be developed utilizing survey data from the Comprehensive Australian Study of Entrepreneurial Emergence (CAUSEE) large-scale longitudinal study of new venture start-ups in Australia. Prior to commencing case studies, 6 scoping interviews were conducted with key stakeholders including industry members, established businesses and government to ensure practical relevance in case development. The venture is considered the unit of analysis with the key informant being the entrepreneur and other management team members where appropriate. Triangulation techniques are used in this research including semi-structured interviews, survey data, onsite visits and secondary documentation website analysis, resumes, and business plans. These 6 sustainability ventures have been selected based on different environmental dynamism conditions including a traditionally mature market (building industry) and a more dynamic, evolving industry (renewable energy/solar ventures). In evaluating multidisciplinary literature, we expect the following external constraints are critical including: technology constraints (seen through lock-in of incumbents existing technology), institutional regulation and standards, access to markets, knowledge and training to nascent and young venture bricolage processes. The case studies will investigate internal constraints including resource fungability, resource combination capabilities, translating complex science/engineering knowledge into salient, valuable market propositions, i.e. appropriate market outcomes, and leveraging relationships may further influence bricolage decisions. ---------- Results and Implications: Intended ventures have been identified within the CAUSEE sample and have agreed to participate and secondary data collection for triangulation purposes has already commenced. Data collection of the case studies commenced 27th of May 2009. Analysis is expected to be completed finalised by 25th September 2009. This paper will report on the pattern of resource constraints and its impact on bricolage behaviours: its subsequent impact on resource deployment within venture creation and venture growth. As such, this research extends the theory of bricolage through the systematic analysis of constraints on resource management processes in sustainability ventures. For practice, this research may assist in providing a better understanding of the resource requirements and processes needed for continued venture persistence and growth in sustainability ventures. In these times of economic uncertainty, a better understanding of the influence on constraints and bricolage: the interplay of behaviours, processes and outcomes may enable greater venture continuance and success.

Truth, obligation and ethics in the propagation of democracy

In this paper I present an analysis of the language used by the National Endowment for Democracy (NED) on its website (NED, 2008). The specific focus of the analysis is on the NED's high usage of the word “should” revealed in computer assisted corpus analysis using Leximancer. Typically we use the word “should” as a term to propose specific courses of action for ourselves and others. It is a marker of obligation and “oughtness”. In other words, its systematic institutional use can be read as a statement of ethics, of how the NED thinks the world ought to behave. As an ostensibly democracy-promoting institution, and one with a clear agenda of implementing American foreign policy, the ethics of NED are worth understanding. Analysis reveals a pattern of grammatical metaphor in which “should” is often deployed counter intuitively, and sometimes ambiguously, as a truth-making tool rather than one for proposing action. The effect is to present NED's imperatives for action as matters of fact rather than ethical or obligatory claims.

Business on-line? An empirical study of factors leading to the adoption of Internet technologies by Australian SMEs

E-commerce technologies such as a website, email and the use of web browsers enables access to large amounts of information, facilitates communication and provides niche companies with an effective mechanism for competing with larger organisations world-wide. However recent literature has shown Australian SMEs have been slow in the uptake of these technologies. The aim of this research was to determine which factors were important in impacting on small firms' decision making in respect of information technology and e-commerce adoption. Findings indicate that generally the more a firm was concerned about its competitive position such a firm was likely to develop a web site. Moreover the 'Industry and Skill Demands' dimension suggested that as the formal education of the owner/manager increased, coupled with the likelihood that the firm was in the transport and storage or communication services industries, and realising the cost of IT adoption was in effect an investment, then such a firm would be inclined to develop a web site.

A national evaluation of the Australian Occupational Therapy Competency Standards (1994): a multistakeholder perspective

This paper summarises results from an evaluation of the adequacy and utility of the Australian Competency Standards for Entry-Level Occupational Therapists © (OT AUSTRALIA, 1994a). It comprised a two-part study, incorporating an online survey of key national stakeholders (n = 26), and 13 focus groups (n = 152) conducted throughout Australia with occupational therapy clinicians, academics, OT AUSTRALIA association and Occupational Therapy Registration Board representatives, as well as university program accreditors. The key recommendations were that: (i) urgent revision to reflect contemporary practice, paradigms, approaches and frameworks is required; (ii) the standards should exemplify basic competence at graduation (not within two years following); (iii) a revision cycle of five years is required; (iv) the Australian Qualifications Framework should be retained, preceded by an introduction describing the scope and nature of occupational therapy practice in the national context; (v) access to the standards should be free and unrestricted to occupational therapists, students and the public via the OT AUSTRALIA (national) website; (vi) the standards should incorporate a succinct executive summary and additional tools or templates formatted to enable occupational therapists to develop professional portfolios and create working documents specific to their workplace; and (vii) language must accommodate contextual variation while striking an appropriate balance between providing instruction and encouraging innovation in practice.

Algebraic analysis of small scale LEX-BES

This paper examines the algebraic cryptanalysis of small scale variants of the LEX-BES. LEX-BES is a stream cipher based on the Advanced Encryption Standard (AES) block cipher. LEX is a generic method proposed for constructing a stream cipher from a block cipher, initially introduced by Biryukov at eSTREAM, the ECRYPT Stream Cipher project in 2005. The Big Encryption System (BES) is a block cipher introduced at CRYPTO 2002 which facilitates the algebraic analysis of the AES block cipher. In this paper, experiments were conducted to find solution of the equation system describing small scale LEX-BES using Gröbner Basis computations. This follows a similar approach to the work by Cid, Murphy and Robshaw at FSE 2005 that investigated algebraic cryptanalysis on small scale variants of the BES. The difference between LEX-BES and BES is that due to the way the keystream is extracted, the number of unknowns in LEX-BES equations is fewer than the number in BES. As far as the author knows, this attempt is the first at creating solvable equation systems for stream ciphers based on the LEX method using Gröbner Basis computations.

Rural Poultry in the Developing world

In 2007 I travelled to Mozambique and Tanzania to photographically document the work undertaken by a group of Australians who are working to bring a self sufficient lifestyle back to the HIV Aids stricken communities of Africa. University of Queensland veterinary researchers have developed a vaccination that can eradicate disease from local rural poultry. The Kyeema Foundation is working in country to supply this vaccine to families battling HIV AIDS and to teach local residents how to successfully implement it. QUT, AusAID and the Kyeema Foundation funded the trip.

Analysis of linear relationships in block ciphers

This thesis is devoted to the study of linear relationships in symmetric block ciphers. A block cipher is designed so that the ciphertext is produced as a nonlinear function of the plaintext and secret master key. However, linear relationships within the cipher can still exist if the texts and components of the cipher are manipulated in a number of ways, as shown in this thesis. There are four main contributions of this thesis. The first contribution is the extension of the applicability of integral attacks from word-based to bitbased block ciphers. Integral attacks exploit the linear relationship between texts at intermediate stages of encryption. This relationship can be used to recover subkey bits in a key recovery attack. In principle, integral attacks can be applied to bit-based block ciphers. However, specific tools to define the attack on these ciphers are not available. This problem is addressed in this thesis by introducing a refined set of notations to describe the attack. The bit patternbased integral attack is successfully demonstrated on reduced-round variants of the block ciphers Noekeon, Present and Serpent. The second contribution is the discovery of a very small system of equations that describe the LEX-AES stream cipher. LEX-AES is based heavily on the 128-bit-key (16-byte) Advanced Encryption Standard (AES) block cipher. In one instance, the system contains 21 equations and 17 unknown bytes. This is very close to the upper limit for an exhaustive key search, which is 16 bytes. One only needs to acquire 36 bytes of keystream to generate the equations. Therefore, the security of this cipher depends on the difficulty of solving this small system of equations. The third contribution is the proposal of an alternative method to measure diffusion in the linear transformation of Substitution-Permutation-Network (SPN) block ciphers. Currently, the branch number is widely used for this purpose. It is useful for estimating the possible success of differential and linear attacks on a particular SPN cipher. However, the measure does not give information on the number of input bits that are left unchanged by the transformation when producing the output bits. The new measure introduced in this thesis is intended to complement the current branch number technique. The measure is based on fixed points and simple linear relationships between the input and output words of the linear transformation. The measure represents the average fraction of input words to a linear diffusion transformation that are not effectively changed by the transformation. This measure is applied to the block ciphers AES, ARIA, Serpent and Present. It is shown that except for Serpent, the linear transformations used in the block ciphers examined do not behave as expected for a random linear transformation. The fourth contribution is the identification of linear paths in the nonlinear round function of the SMS4 block cipher. The SMS4 block cipher is used as a standard in the Chinese Wireless LAN Wired Authentication and Privacy Infrastructure (WAPI) and hence, the round function should exhibit a high level of nonlinearity. However, the findings in this thesis on the existence of linear relationships show that this is not the case. It is shown that in some exceptional cases, the first four rounds of SMS4 are effectively linear. In these cases, the effective number of rounds for SMS4 is reduced by four, from 32 to 28. The findings raise questions about the security provided by SMS4, and might provide clues on the existence of a flaw in the design of the cipher.

A story worth telling : putting oral history and digital collections online in cultural institutions

Digital platforms in cultural institutions offer exciting opportunities for oral history and digital storytelling that can augment and enrich traditional collections. The way in which cultural institutions allow access to the public is changing dramatically, prompting substantial expansions of their oral history and digital story holdings. In Queensland, Australia, public libraries and museums are becoming innovative hubs of a wide assortment of collections that represent a cross-section of community groups and organisations through the integration of oral history and digital storytelling. The State Library of Queensland (SLQ) features digital stories online to encourage users to explore what the institution has in the catalogue through their website. Now SLQ also offers oral history interviews online, to introduce users to oral history and other components of their collections,- such as photographs and documents to current, as well as new users. This includes the various departments, Indigenous centres and regional libraries affiliated with SLQ statewide, who are often unable to access the materials held within, or even full information about, the collections available within the institution. There has been a growing demand for resources and services that help to satisfy community enthusiasm and promote engagement. Demand increases as public access to affordable digital media technologies increases, and as community or marginalised groups become interested in do it yourself (DIY) history; and SLQ encourages this. This paper draws on the oral history and digital story-based research undertaken by the Queensland University of Technology (QUT) for the State Library of Queensland including: the Apology Collection: The Prime Minister’s apology to Australia’s Indigenous Stolen Generation; Five Senses: regional Queensland artists; Gay history of Brisbane; and The Queensland Business Leaders Hall of Fame.

Algebraic analysis of small Scale LEX-BES

This work examines the algebraic cryptanalysis of small scale variants of the LEX-BES. LEX-BES is a stream cipher based on the Advanced Encryption Standard (AES) block cipher. LEX is a generic method proposed for constructing a stream cipher from a block cipher, initially introduced by Biryukov at eSTREAM, the ECRYPT Stream Cipher project in 2005. The Big Encryption System (BES) is a block cipher introduced at CRYPTO 2002 which facilitates the algebraic analysis of the AES block cipher. In this article, experiments were conducted to find solutions of equation systems describing small scale LEX-BES using Gröbner Basis computations. This follows a similar approach to the work by Cid, Murphy and Robshaw at FSE 2005 that investigated algebraic cryptanalysis on small scale variants of the BES. The difference between LEX-BES and BES is that due to the way the keystream is extracted, the number of unknowns in LEX-BES equations is fewer than the number in BES. As far as the authors know, this attempt is the first at creating solvable equation systems for stream ciphers based on the LEX method using Gröbner Basis computations.

Power electronics education electronic book (PEEEB)

In this website, you can virtually attend all lectures, tutorials, computer Labs and quizzes and also access to lecture notes.

Linearity within the SMS4 Block Cipher

We present several new observations on the SMS4 block cipher, and discuss their cryptographic significance. The crucial observation is the existence of fixed points and also of simple linear relationships between the bits of the input and output words for each component of the round functions for some input words. This implies that the non-linear function T of SMS4 does not appear random and that the linear transformation provides poor diffusion. Furthermore, the branch number of the linear transformation in the key scheduling algorithm is shown to be less than optimal. The main security implication of these observations is that the round function is not always non-linear. Due to this linearity, it is possible to reduce the number of effective rounds of SMS4 by four. We also investigate the susceptibility of SMS4 to further cryptanalysis. Finally, we demonstrate a successful differential attack on a slightly modified variant of SMS4. These findings raise serious questions on the security provided by SMS4.

QUT Digital Repository: Integrating the University’s IR with other digital collections

Queensland University of Technology’s Institutional Repository, QUT ePrints (http://eprints.qut.edu.au/), was established in 2003. With the help of an institutional mandate (endorsed in 2004) the repository now holds over 11,000 open access publications. The repository’s success is celebrated within the University and acknowledged nationally and internationally. QUT ePrints was built on GNU EPrints open source repository software (currently running v.3.1.3) and was originally configured to accommodate open access versions of the traditional range of research publications (journal articles, conference papers, books, book chapters and working papers). However, in 2009, the repository’s scope, content and systems were broadened and the ‘QUT Digital repository’ is now a service encompassing a range of digital collections, services and systems. For a work to be accepted in to the institutional repository, at least one of the authors/creators must have a current affiliation with QUT. However, the success of QUT ePrints in terms of its capacity to increase the visibility and accessibility of our researchers' scholarly works resulted in requests to accept digital collections of works which were out of scope. To address this need, a number of parallel digital collections have been developed. These collections include, OZcase, a collection of legal research materials and ‘The Sugar Industry Collection’; a digitsed collection of books and articles on sugar cane production and processing. Additionally, the Library has responded to requests from academics for a service to support the publication of new, and existing, peer reviewed open access journals. A project is currently underway to help a group of senior QUT academics publish a new international peer reviewed journal. The QUT Digital Repository website will be a portal for access to a range of resources to support copyright management. It is likely that it will provide an access point for the institution’s data repository. The data repository, provisionally named the ‘QUT Data Commons’, is currently a work-in-progress. The metadata for some QUT datasets will also be harvested by and discoverable via ‘Research Data Australia’, the dataset discovery service managed by the Australian National Data Service (ANDS). QUT Digital repository will integrate a range of technologies and services related to scholarly communication. This paper will discuss the development of the QUT Digital Repository, its strategic functions, the stakeholders involved and lessons learned.