The transformation of banana with potential virus resistance genes

One approach to reducing the yield losses caused by banana viral diseases is the use of genetic engineering and pathogen-derived resistance strategies to generate resistant cultivars. The development of transgenic virus resistance requires an efficient banana transformation method, particularly for commercially important 'Cavendish' type cultivars such as 'Grand Nain'. Prior to this study, only two examples of the stable transformation of banana had been reported, both of which demonstrated the principle of transformation but did not characterise transgenic plants in terms of the efficiency at which individual transgenic lines were generated, relative activities of promoters in stably transformed plants, and the stability of transgene expression. The aim of this study was to develop more efficient transformation methods for banana, assess the activity of some commonly used and also novel promoters in stably transformed plants, and transform banana with genes that could potentially confer resistance to banana bunchy top nanovirus (BBTV) and banana bract mosaic potyvirus (BBrMV). A regeneration system using immature male flowers as the explant was established. The frequency of somatic embryogenesis in male flower explants was influenced by the season in which the inflorescences were harvested. Further, the media requirements of various banana cultivars in respect to the 2,4-D concentration in the initiation media also differed. Following the optimisation of these and other parameters, embryogenic cell suspensions of several banana (Musa spp.) cultivars including 'Grand Nain' (AAA), 'Williams' (AAA), 'SH-3362' (AA), 'Goldfinger' (AAAB) and 'Bluggoe' (ABB) were successfully generated. Highly efficient transformation methods were developed for both 'Bluggoe' and 'Grand Nain'; this is the first report of microprojectile bombardment transformation of the commercially important 'Grand Nain' cultivar. Following bombardment of embryogenic suspension cells, regeneration was monitored from single transfom1ed cells to whole plants using a reporter gene encoding the green fluorescent protein (gfp). Selection with kanamycin enabled the regeneration of a greater number of plants than with geneticin, while still preventing the regeneration of non-transformed plants. Southern hybridisation confirmed the neomycin phosphotransferase gene (npt II) was stably integrated into the banana genome and that multiple transgenic lines were derived from single bombardments. The activity, stability and tissue specificity of the cauliflower mosaic virus 358 (CaMV 35S) and maize polyubiquitin-1 (Ubi-1) promoters were examined. In stably transformed banana, the Ubi-1 promoter provided approximately six-fold higher p-glucuronidase (GUS) activity than the CaMV 35S promoter, and both promoters remained active in glasshouse grown plants for the six months they were observed. The intergenic regions ofBBTV DNA-I to -6 were isolated and fused to either the uidA (GUS) or gfjJ reporter genes to assess their promoter activities. BBTV promoter activity was detected in banana embryogenic cells using the gfp reporter gene. Promoters derived from BBTV DNA-4 and -5 generated the highest levels of transient activity, which were greater than that generated by the maize Ubi-1 promoter. In transgenic banana plants, the activity of the BBTV DNA-6 promoter (BT6.1) was restricted to the phloem of leaves and roots, stomata and root meristems. The activity of the BT6.1 promoter was enhanced by the inclusion of intron-containing fragments derived from the maize Ubi-1, rice Act-1, and sugarcane rbcS 5' untranslated regions in GUS reporter gene constructs. In transient assays in banana, the rice Act-1 and maize Ubi-1 introns provided the most significant enhancement, increasing expression levels 300-fold and 100-fold, respectively. The sugarcane rbcS intron increased expression about 10-fold. In stably transformed banana plants, the maize Ubi-1 intron enhanced BT6.1 promoter activity to levels similar to that of the CaMV 35S promoter, but did not appear to alter the tissue specificity of the promoter. Both 'Grand Nain' and 'Bluggoe' were transformed with constructs that could potentially confer resistance to BBTV and BBrMV, including constructs containing BBTV DNA-1 major and internal genes, BBTV DNA-5 gene, and the BBrMV coat protein-coding region all under the control of the Ubi-1 promoter, while the BT6 promoter was used to drive the npt II selectable marker gene. At least 30 transgenic lines containing each construct were identified and replicates of each line are currently being generated by micropropagation in preparation for virus challenge.

The development of a serological-based diagnostic test for Dasheen mosaic potyvirus (DsMV)

Dasheen mosaic potyvirus (DsMV) is an important virus affecting taro. The virus has been found wherever taro is grown and infects both the edible and ornamental aroids, causing yield losses of up to 60%. The presence of DsMV, and other viruses,prevents the international movement of taro germplasm between countries. This has a significant negative impact on taro production in many countries due to the inability to access improved taro lines produced in breeding programs. To overcome this problem, sensitive and reliable virus diagnostic tests need to be developed to enable the indexing of taro germplasm. The aim of this study was to generate an antiserum against a recombinant DsMV coat protein (CP) and to develop a serological-based diagnostic test that would detect Pacific Island isolates of the virus. The CP-coding region of 16 DsMV isolates from Papua New Guinea, Samoa, Solomon Islands, French Polynesia, New Caledonia and Vietnam were amplified,cloned and sequenced. The size of the CP-coding region ranged from 939 to 1038 nucleotides and encoded putative proteins ranged from 313 to 346 amino acids, with the molecular mass ranging from 34 to 38 kDa. Analysis ofthe amino acid sequences revealed the presence of several amino acid motifs typically found in potyviruses,including DAG, WCIE/DN, RQ and AFDF. When the amino acid sequences were compared with each other and the DsMV sequences on the database, the maximum variability was21.9%. When the core region ofthe CP was analysed, the maximum variability dropped to 6% indicating most variability was present in the N terminus. Within seven PNG isolates ofDsMV, the maximum variability was 16.9% and 3.9% over the entire CP-coding region and core region, respectively. The sequence ofPNG isolate P1 was most similar to all other sequences. Phylogenetic analysis indicated that almost all isolates grouped according to their provenance. Further, the seven PNG isolates were grouped according to the region within PNG from which they were obtained. Due to the extensive variability over the entire CP-coding region, the core region ofthe CP ofPNG isolate Pl was cloned into a protein expression vector and expressed as a recombinant protein. The protein was purified by chromatography and SDS-PAGE and used as an antigen to generate antiserum in a rabbit. In western blots, the antiserum reacted with bands of approximately 45-47 kDa in extracts from purified DsMV and from known DsMV -infected plants from PNG; no bands were observed using healthy plant extracts. The antiserum was subsequently incorporated into an indirect ELISA. This procedure was found to be very sensitive and detected DsMV in sap diluted at least 1:1,000. Using both western blot and ELISA formats,the antiserum was able to detect a wide range ofDsMV isolates including those from Australia, New Zealand, Fiji, French Polynesia, New Caledonia, Papua New Guinea, Samoa, Solomon Islands and Vanuatu. These plants were verified to be infected with DsMV by RT-PCR. In specificity tests, the antiserum was also found to react with sap from plants infected with SCMV, PRSV-P, PRSV-W, but not with PVY or CMV -infected plants.

aa-cell in practice : an approach to musical live coding

Live coding performances provide a context with particular demands and limitations for music making. In this paper we discuss how as the live coding duo aa-cell we have responded to these challenges, and what this experience has revealed about the computational representation of music and approaches to interactive computer music performance. In particular we have identified several effective and efficient processes that underpin our practice including probability, linearity, periodicity, set theory, and recursion and describe how these are applied and combined to build sophisticated musical structures. In addition, we outline aspects of our performance practice that respond to the improvisational, collaborative and communicative requirements of musical live coding.

Machine learning techniques to improve software quality

A significant proportion of the cost of software development is due to software testing and maintenance. This is in part the result of the inevitable imperfections due to human error, lack of quality during the design and coding of software, and the increasing need to reduce faults to improve customer satisfaction in a competitive marketplace. Given the cost and importance of removing errors improvements in fault detection and removal can be of significant benefit. The earlier in the development process faults can be found, the less it costs to correct them and the less likely other faults are to develop. This research aims to make the testing process more efficient and effective by identifying those software modules most likely to contain faults, allowing testing efforts to be carefully targeted. This is done with the use of machine learning algorithms which use examples of fault prone and not fault prone modules to develop predictive models of quality. In order to learn the numerical mapping between module and classification, a module is represented in terms of software metrics. A difficulty in this sort of problem is sourcing software engineering data of adequate quality. In this work, data is obtained from two sources, the NASA Metrics Data Program, and the open source Eclipse project. Feature selection before learning is applied, and in this area a number of different feature selection methods are applied to find which work best. Two machine learning algorithms are applied to the data - Naive Bayes and the Support Vector Machine - and predictive results are compared to those of previous efforts and found to be superior on selected data sets and comparable on others. In addition, a new classification method is proposed, Rank Sum, in which a ranking abstraction is laid over bin densities for each class, and a classification is determined based on the sum of ranks over features. A novel extension of this method is also described based on an observed polarising of points by class when rank sum is applied to training data to convert it into 2D rank sum space. SVM is applied to this transformed data to produce models the parameters of which can be set according to trade-off curves to obtain a particular performance trade-off.

Towards learner autonomy : an assessment for learning approach

Using Assessment for Learning (AfL) may develop learner autonomy however, very often AfL is reduced to a set of strategies that do not always achieve the desired outcome. This research adopted a different approach that examined AfL as a cultural practice, situated within influential social relationships that shape learner identity. The study addressed the question “What are the qualities of the teacher-student relationship that support student learning autonomy in an AfL context?” Three case studies of the interactions of Queensland middle school teachers and their classes of Year 7, 8 and 9 were developed over one year. Data were collected from field notes and video recordings of classroom interactions and individual and focus group interviews with teachers and students. The analysis began with a close look at the field data. Interpretations that emerged from a sociocultural theoretical understanding were helpful in informing the process of analysis. Themes and patterns of interrelationships were identified through thematic coding using a constant comparative approach. Validation was achieved through methodological triangulation. Four findings that inform an understanding of AfL and the development of learner autonomy emerged. Firstly, autonomy is theorised as a context-specific identity mediated through the teacher-student relationship. Secondly, it was observed that learners negotiated their identities as knowers through AfL practices in various tacit, explicit, group and individual ways in a ‘generative dance’ of knowing in action (Cook & Brown, 2005). Thirdly, teachers and learners negotiated their participation by drawing from identities in multiple communities of practice. Finally it is proposed that a new participative identity or narrative for assessment is needed. This study contributes to understandings about teacher AfL practices that can help build teacher assessment capacity. Importantly, autonomy is understood as an identity that is available to all learners. This study is also significant as it affirms the importance of teacher assessment to support learners in developing autonomy, a focus that challenges the singular assessment policy focus on measuring performance. Finally this study contributes to a sociocultural theoretical understanding of AfL.

An exploration of the technical, social and macro-societal pressures influencing the adoption of online social networking services by organisations

Measuring the business value that Internet technologies deliver for organisations has proven to be a difficult and elusive task, given their complexity and increased embeddedness within the value chain. Yet, despite the lack of empirical evidence that links the adoption of Information Technology (IT) with increased financial performance, many organisations continue to adopt new technologies at a rapid rate. This is evident in the widespread adoption of Web 2.0 online Social Networking Services (SNSs) such as Facebook, Twitter and YouTube. These new Internet based technologies, widely used for social purposes, are being employed by organisations to enhance their business communication processes. However, their use is yet to be correlated with an increase in business performance. Owing to the conflicting empirical evidence that links prior IT applications with increased business performance, IT, Information Systems (IS), and E-Business Model (EBM) research has increasingly looked to broader social and environmental factors as a means for examining and understanding the broader influences shaping IT, IS and E-Business (EB) adoption behaviour. Findings from these studies suggest that organisations adopt new technologies as a result of strong external pressures, rather than a clear measure of enhanced business value. In order to ascertain if this is the case with the adoption of SNSs, this study explores how organisations are creating value (and measuring that value) with the use of SNSs for business purposes, and the external pressures influencing their adoption. In doing so, it seeks to address two research questions: 1. What are the external pressures influencing organisations to adopt SNSs for business communication purposes? 2. Are SNSs providing increased business value for organisations, and if so, how is that value being captured and measured? Informed by the background literature fields of IT, IS, EBM, and Web 2.0, a three-tiered theoretical framework is developed that combines macro-societal, social and technological perspectives as possible causal mechanisms influencing the SNS adoption event. The macro societal view draws on the concept of Castells. (1996) network society and the behaviour of crowds, herds and swarms, to formulate a new explanatory concept of the network vortex. The social perspective draws on key components of institutional theory (DiMaggio & Powell, 1983, 1991), and the technical view draws from the organising vision concept developed by Swanson and Ramiller (1997). The study takes a critical realist approach, and conducts four stages of data collection and one stage of data coding and analysis. Stage 1 consisted of content analysis of websites and SNSs of many organisations, to identify the types of business purposes SNSs are being used for. Stage 2 also involved content analysis of organisational websites, in order to identify suitable sample organisations in which to conduct telephone interviews. Stage 3 consisted of conducting 18 in-depth, semi-structured telephone interviews within eight Australian organisations from the Media/Publishing and Galleries, Libraries, Archives and Museum (GLAM) industries. These sample organisations were considered leaders in the use of SNSs technologies. Stage 4 involved an SNS activity count of the organisations interviewed in Stage 3, in order to rate them as either Advanced Innovator (AI) organisations, or Learning Focussed (LF) organisations. A fifth stage of data coding and analysis of all four data collection stages was conducted, based on the theoretical framework developed for the study, and using QSR NVivo 8 software. The findings from this study reveal that SNSs have been adopted by organisations for the purpose of increasing business value, and as a result of strong social and macro-societal pressures. SNSs offer organisations a wide range of value enhancing opportunities that have broader benefits for customers and society. However, measuring the increased business value is difficult with traditional Return On Investment (ROI) mechanisms, ascertaining the need for new value capture and measurement rationales, to support the accountability of SNS adoption practices. The study also identified the presence of technical, social and macro-societal pressures, all of which influenced SNS adoption by organisations. These findings contribute important theoretical insight into the increased complexity of pressures influencing technology adoption rationales by organisations, and have important practical implications for practice, by reflecting the expanded global online networks in which organisations now operate. The limitations of the study include the small number of sample organisations in which interviews were conducted, its limited generalisability, and the small range of SNSs selected for the study. However, these were compensated in part by the expertise of the interviewees, and the global significance of the SNSs that were chosen. Future research could replicate the study to a larger sample from different industries, sectors and countries. It could also explore the life cycle of SNSs in a longitudinal study, and map how the technical, social and macro-societal pressures are emphasised through stages of the life cycle. The theoretical framework could also be applied to other social fad technology adoption studies.

Understanding the phenomenon of IS success in China through the lens of the IS-Impact model

This study is conducted within the IS-Impact Research Track at Queensland University of Technology (QUT). The goal of the IS-Impact Track is, “to develop the most widely employed model for benchmarking information systems in organizations for the joint benefit of both research and practice” (Gable et al, 2006). IS-Impact is defined as “a measure at a point in time, of the stream of net benefits from the IS, to date and anticipated, as perceived by all key-user-groups” (Gable Sedera and Chan, 2008). Track efforts have yielded the bicameral IS-Impact measurement model; the “impact” half includes Organizational-Impact and Individual-Impact dimensions; the “quality” half includes System-Quality and Information-Quality dimensions. The IS-Impact model, by design, is intended to be robust, simple and generalizable, to yield results that are comparable across time, stakeholders, different systems and system contexts. The model and measurement approach employ perceptual measures and an instrument that is relevant to key stakeholder groups, thereby enabling the combination or comparison of stakeholder perspectives. Such a validated and widely accepted IS-Impact measurement model has both academic and practical value. It facilitates systematic operationalization of a main dependent variable in research (IS-Impact), which can also serve as an important independent variable. For IS management practice it provides a means to benchmark and track the performance of information systems in use. The objective of this study is to develop a Mandarin version IS-Impact model, encompassing a list of China-specific IS-Impact measures, aiding in a better understanding of the IS-Impact phenomenon in a Chinese organizational context. The IS-Impact model provides a much needed theoretical guidance for this investigation of ES and ES impacts in a Chinese context. The appropriateness and soundness of employing the IS-Impact model as a theoretical foundation are evident: the model originated from a sound theory of IS Success (1992), developed through rigorous validation, and also derived in the context of Enterprise Systems. Based on the IS-Impact model, this study investigates a number of research questions (RQs). Firstly, the research investigated what essential impacts have been derived from ES by Chinese users and organizations [RQ1]. Secondly, we investigate which salient quality features of ES are perceived by Chinese users [RQ2]. Thirdly, we seek to answer whether the quality and impacts measures are sufficient to assess ES-success in general [RQ3]. Lastly, the study attempts to address whether the IS-Impact measurement model is appropriate for Chinese organizations in terms of evaluating their ES [RQ4]. An open-ended, qualitative identification survey was employed in the study. A large body of short text data was gathered from 144 Chinese users and 633 valid IS-Impact statements were generated from the data set. A generally inductive approach was applied in the qualitative data analysis. Rigorous qualitative data coding resulted in 50 first-order categories with 6 second-order categories that were grounded from the context of Chinese organization. The six second-order categories are: 1) System Quality; 2) Information Quality; 3) Individual Impacts;4) Organizational Impacts; 5) User Quality and 6) IS Support Quality. The final research finding of the study is the contextualized Mandarin version IS-Impact measurement model that includes 38 measures organized into 4 dimensions: System Quality, information Quality, Individual Impacts and Organizational Impacts. The study also proposed two conceptual models to harmonize the IS-Impact model and the two emergent constructs – User Quality and IS Support Quality by drawing on previous IS effectiveness literatures and the Work System theory proposed by Alter (1999) respectively. The study is significant as it is the first effort that empirically and comprehensively investigates IS-Impact in China. Specifically, the research contributions can be classified into theoretical contributions and practical contributions. From the theoretical perspective, through qualitative evidence, the study test and consolidate IS-Impact measurement model in terms of the quality of robustness, completeness and generalizability. The unconventional research design exhibits creativity of the study. The theoretical model does not work as a top-down a priori seeking for evidence demonstrating its credibility; rather, the study allows a competitive model to emerge from the bottom-up and open-coding analysis. Besides, the study is an example extending and localizing pre-existing theory developed in Western context when the theory is introduced to a different context. On the other hand, from the practical perspective, It is first time to introduce prominent research findings in field of IS Success to Chinese academia and practitioner. This study provides a guideline for Chinese organizations to assess their Enterprise System, and leveraging IT investment in the future. As a research effort in ITPS track, this study contributes the research team with an alternative operationalization of the dependent variable. The future research can take on the contextualized Mandarin version IS-Impact framework as a theoretical a priori model, further quantitative and empirical testing its validity.

Errata supplement for QUT thesis 'Heavy vehicle suspensions : testing and analysis'

Errata supplement to QUT thesis: 'Heavy vehicle suspensions : testing and analysis'

Comparative study of coagulase negative Staphylococci (CoNS) from clinical isolates, skin and nasal sources

Staphylococci are important pathogenic bacteria responsible for a range of diseases in humans. The most frequently isolated microorganisms in a hospital microbiology laboratory are staphylococci. The general classification of staphylococci divides them into two major groups; Coagulase-positive staphylococci (e.g. Staphylococcus aureus) and Coagulase-negative staphylococci (e.g. Staphylococcus epidermidis). Coagulase-negative staphylococcal (CoNS) isolates include a variety of species and many different strains but are often dominated by the most important organism of this group, S. epidermidis. Currently, these organisms are regarded as important pathogenic organisms causing infections related to prosthetic materials and surgical wounds. A significant number of S. epidermidis isolates are also resistant to different antimicrobial agents. Virulence factors in CoNS are not very clearly established and not well documented. S. epidermidis is evolving as a resistant and powerful microbe related to nosocomial infections because it has different properties which independently, and in combination, make it a successful infectious agent, especially in the hospital environment. Such characteristics include biofilm formation, drug resistance and the evolution of genetic variables. The purpose of this project was to develop a novel SNP genotyping method to genotype S. epidermidis strains originating from hospital patients and healthy individuals. High-Resolution Melt Analysis was used to assign binary typing profiles to both clinical and commensal strains using a new bioinformatics approach. The presence of antibiotic resistance genes and biofilm coding genes were also interrogated in these isolates.

Defining the role of phytoene synthase in carotenoid accumulation of high provitamin A bananas

Vitamin A deficiency (VAD) is a serious problem in developing countries, affecting approximately 127 million children of preschool age and 7.2 million pregnant women each year. However, this deficiency is readily treated and prevented through adequate nutrition. This can potentially be achieved through genetically engineered biofortification of staple food crops to enhance provitamin A (pVA) carotenoid content. Bananas are the fourth most important food crop with an annual production of 100 million tonnes and are widely consumed in areas affected by VAD. However, the fruit pVA content of most widely consumed banana cultivars is low (~ 0.2 to 0.5 ìg/g dry weight). This includes cultivars such as the East African highland banana (EAHB), the staple crop in countries such as Uganda, where annual banana consumption is approximately 250 kg per person. This fact, in addition to the agronomic properties of staple banana cultivars such as vegetative reproduction and continuous cropping, make bananas an ideal target for pVA enhancement through genetic engineering. Interestingly, there are banana varieties known with high fruit pVA content (up to 27.8 ìg/g dry weight), although they are not widely consumed due to factors such as cultural preference and availability. The genes involved in carotenoid accumulation during banana fruit ripening have not been well studied and an understanding of the molecular basis for the differential capacity of bananas to accumulate carotenoids may impact on the effective production of genetically engineered high pVA bananas. The production of phytoene by the enzyme phytoene synthase (PSY) has been shown to be an important rate limiting determinant of pVA accumulation in crop systems such as maize and rice. Manipulation of this gene in rice has been used successfully to produce Golden Rice, which exhibits higher seed endosperm pVA levels than wild type plants. Therefore, it was hypothesised that differences between high and low pVA accumulating bananas could be due either to differences in PSY enzyme activity or factors regulating the expression of the psy gene. Therefore, the aim of this thesis was to investigate the role of PSY in accumulation of pVA in banana fruit of representative high (Asupina) and low (Cavendish) pVA banana cultivars by comparing the nucleic acid and encoded amino acid sequences of the banana psy genes, in vivo enzyme activity of PSY in rice callus and expression of PSY through analysis of promoter activity and mRNA levels. Initially, partial sequences of the psy coding region from five banana cultivars were obtained using reverse transcriptase (RT)-PCR with degenerate primers designed to conserved amino acids in the coding region of available psy sequences from other plants. Based on phylogenetic analysis and comparison to maize psy sequences, it was found that in banana, psy occurs as a gene family of at least three members (psy1, psy2a and psy2b). Subsequent analysis of the complete coding regions of these genes from Asupina and Cavendish suggested that they were all capable of producing functional proteins due to high conservation in the catalytic domain. However, inability to obtain the complete mRNA sequences of Cavendish psy2a, and isolation of two non-functional Cavendish psy2a coding region variants, suggested that psy2a expression may be impaired in Cavendish. Sequence analysis indicated that these Cavendish psy2a coding region variants may have resulted from alternate splicing. Evidence of alternate splicing was also observed in one Asupina psy1 coding region variant, which was predicted to produce a functional PSY1 isoform. The complete mRNA sequence of the psy2b coding regions could not be isolated from either cultivar. Interestingly, psy1 was cloned predominantly from leaf while psy2 was obtained preferentially from fruit, suggesting some level of tissue-specific expression. The Asupina and Cavendish psy1 and psy2a coding regions were subsequently expressed in rice callus and the activity of the enzymes compared in vivo through visual observation and quantitative measurement of carotenoid accumulation. The maize B73 psy1 coding region was included as a positive control. After several weeks on selection, regenerating calli showed a range of colours from white to dark orange representing various levels of carotenoid accumulation. These results confirmed that the banana psy coding regions were all capable of producing functional enzymes. No statistically significant differences in levels of activity were observed between banana PSYs, suggesting that differences in PSY activity were not responsible for differences in the fruit pVA content of Asupina and Cavendish. The psy1 and psy2a promoter sequences were isolated from Asupina and Cavendish gDNA using a PCR-based genome walking strategy. Interestingly, three Cavendish psy2a promoter clones of different sizes, representing possible allelic variants, were identified while only single promoter sequences were obtained for the other Asupina and Cavendish psy genes. Bioinformatic analysis of these sequences identified motifs that were previously characterised in the Arabidopsis psy promoter. Notably, an ATCTA motif associated with basal expression in Arabidopsis was identified in all promoters with the exception of two of the Cavendish psy2a promoter clones (Cpsy2apr2 and Cpsy2apr3). G1 and G2 motifs, linked to light-regulated responses in Arabidopsis, appeared to be differentially distributed between psy1 and psy2a promoters. In the untranscribed regulatory regions, the G1 motifs were found only in psy1 promoters, while the G2 motifs were found only in psy2a. Interestingly, both ATCTA and G2 motifs were identified in the 5’ UTRs of Asupina and Cavendish psy1. Consistent with other monocot promoters, introns were present in the Asupina and Cavendish psy1 5’ UTRs, while none were observed in the psy2a 5’ UTRs. Promoters were cloned into expression constructs, driving the â-glucuronidase (GUS) reporter gene. Transient expression of the Asupina and Cavendish psy1 and psy2a promoters in both Cavendish embryogenic cells and Cavendish fruit demonstrated that all promoters were active, except Cpsy2apr2 and Cpsy2apr3. The functional Cavendish psy2a promoter (Cpsy2apr1) appeared to have activity similar to the Asupina psy2a promoter. The activities of the Asupina and Cavendish psy1 promoters were similar to each other, and comparable to those of the functional psy2a promoters. Semi-quantitative PCR analysis of Asupina and Cavendish psy1 and psy2a transcripts showed that psy2a levels were high in green fruit and decreased during ripening, reinforcing the hypothesis that fruit pVA levels were largely dependent on levels of psy2a expression. Additionally, semi-quantitative PCR using intron-spanning primers indicated that high levels of unprocessed psy2a and psy2b mRNA were present in the ripe fruit of Cavendish but not in Asupina. This raised the possibility that differences in intron processing may influence pVA accumulation in Asupina and Cavendish. In this study the role of PSY in banana pVA accumulation was analysed at a number of different levels. Both mRNA accumulation and promoter activity of psy genes studied were very similar between Asupina and Cavendish. However, in several experiments there was evidence of cryptic or alternate splicing that differed in Cavendish compared to Asupina, although these differences were not conclusively linked to the differences in fruit pVA accumulation between Asupina and Cavendish. Therefore, other carotenoid biosynthetic genes or regulatory mechanisms may be involved in determining pVA levels in these cultivars. This study has contributed to an increased understanding of the role of PSY in the production of pVA carotenoids in banana fruit, corroborating the importance of this enzyme in regulating carotenoid production. Ultimately, this work may serve to inform future research into pVA accumulation in important crop varieties such as the EAHB and the discovery of avenues to improve such crops through genetic modification.

Measuring bitrate and quality trade-off in a fast region-of-interest based video coding

Prevailing video adaptation solutions change the quality of the video uniformly throughout the whole frame in the bitrate adjustment process; while region-of-interest (ROI)-based solutions selectively retains the quality in the areas of the frame where the viewers are more likely to pay more attention to. ROI-based coding can improve perceptual quality and viewer satisfaction while trading off some bandwidth. However, there has been no comprehensive study to measure the bitrate vs. perceptual quality trade-off so far. The paper proposes an ROI detection scheme for videos, which is characterized with low computational complexity and robustness, and measures the bitrate vs. quality trade-off for ROI-based encoding using a state-of-the-art H.264/AVC encoder to justify the viability of this type of encoding method. The results from the subjective quality test reveal that ROI-based encoding achieves a significant perceptual quality improvement over the encoding with uniform quality at the cost of slightly more bits. Based on the bitrate measurements and subjective quality assessments, the bitrate and the perceptual quality estimation models for non-scalable ROI-based video coding (AVC) are developed, which are found to be similar to the models for scalable video coding (SVC).

Voices of experience : opportunities to influence creatively the designing of school libraries

Using a critical ethnographic approach this study investigates the potential for multiple voices of experience, of educators, designers/architects, education facility planners and students/learners, to influence creatively the designing of school libraries. School libraries are considered as social and cultural entities within the contexts of school life and of wider society. It is proposed that school library designing is a social interaction of concern to those influenced by its practices and outcomes. School library designing is therefore of significance to educators and students as well as to those with professionally accredited involvement in school library designing, such as designers/architects and education facility planners. The study contends that current approaches to educational space designing, including school libraries, amplify the voices of accredited designers and diminish or silence the voices of the user participants. The study is conceptualised as creative processes of discovery, through which attention is paid to the voices of experience of user and designer participants, and is concerned with their understandings and experiences of school libraries and their understandings and experiences of designing. Grounded theory coding (Charmaz) is used for initial categorising of interview data. Critical discourse analysis (CDA, Fairclough) is used as analytical tool for reflection on the literature and for analysis of the small stories gathered through semi-structured interviews, field observations and documents. The critical interpretive stance taken through CDA, enables discussions of aspects of power associated with the understandings and experiences of participants, and for recognition of creative possibilities and creative influence within and beyond current conditions. Through an emphasis on prospects for educators and students as makers of the spaces and places of learning, in particular in school libraries, the study has the potential to inform education facility designing practices and design participant relationships, and to contribute more broadly to knowledge in the fields of education, design, architecture, and education facility planning.

Quality metrics for assessing security-critical computer programs

Existing secure software development principles tend to focus on coding vulnerabilities, such as buffer or integer overflows, that apply to individual program statements, or issues associated with the run-time environment, such as component isolation. Here we instead consider software security from the perspective of potential information flow through a program’s object-oriented module structure. In particular, we define a set of quantifiable "security metrics" which allow programmers to quickly and easily assess the overall security of a given source code program or object-oriented design. Although measuring quality attributes of object-oriented programs for properties such as maintainability and performance has been well-covered in the literature, metrics which measure the quality of information security have received little attention. Moreover, existing securityrelevant metrics assess a system either at a very high level, i.e., the whole system, or at a fine level of granularity, i.e., with respect to individual statements. These approaches make it hard and expensive to recognise a secure system from an early stage of development. Instead, our security metrics are based on well-established compositional properties of object-oriented programs (i.e., data encapsulation, cohesion, coupling, composition, extensibility, inheritance and design size), combined with data flow analysis principles that trace potential information flow between high- and low-security system variables. We first define a set of metrics to assess the security quality of a given object-oriented system based on its design artifacts, allowing defects to be detected at an early stage of development. We then extend these metrics to produce a second set applicable to object-oriented program source code. The resulting metrics make it easy to compare the relative security of functionallyequivalent system designs or source code programs so that, for instance, the security of two different revisions of the same system can be compared directly. This capability is further used to study the impact of specific refactoring rules on system security more generally, at both the design and code levels. By measuring the relative security of various programs refactored using different rules, we thus provide guidelines for the safe application of refactoring steps to security-critical programs. Finally, to make it easy and efficient to measure a system design or program’s security, we have also developed a stand-alone software tool which automatically analyses and measures the security of UML designs and Java program code. The tool’s capabilities are demonstrated by applying it to a number of security-critical system designs and Java programs. Notably, the validity of the metrics is demonstrated empirically through measurements that confirm our expectation that program security typically improves as bugs are fixed, but worsens as new functionality is added.