Characterisation and application of a bovine U6 promoter for expression of short hairpin RNAs

Background
The use of small interfering RNA (siRNA) molecules in animals to achieve double-stranded RNA-mediated interference (RNAi) has recently emerged as a powerful method of sequence-specific gene knockdown. As DNA-based expression of short hairpin RNA (shRNA) for RNAi may offer some advantages over chemical and in vitro synthesised siRNA, a number of vectors for expression of shRNA have been developed. These often feature polymerase III (pol. III) promoters of either mouse or human origin.
Results
To develop a shRNA expression vector specifically for bovine RNAi applications, we identified and characterised a novel bovine U6 small nuclear RNA (snRNA) promoter from bovine sequence data. This promoter is the putative bovine homologue of the human U6-8 snRNA promoter, and features a number of functional sequence elements that are characteristic of these types of pol. III promoters. A PCR based cloning strategy was used to incorporate this promoter sequence into plasmid vectors along with shRNA sequences for RNAi. The promoter was then used to express shRNAs, which resulted in the efficient knockdown of an exogenous reporter gene and an endogenous bovine gene.
Conclusion
We have mined data from the bovine genome sequencing project to identify a functional bovine U6 promoter and used the promoter sequence to construct a shRNA expression vector. The use of this native bovine promoter in shRNA expression is an important component of our future development of RNAi therapeutic and transgenic applications in bovine species.

Civilisation, culture and the hybrid self in the work of Robert Ezra Park

Contemporary discussions on hybridity in cultural and ethnic studies have overlooked the work of the Chicago sociologist Robert E. Park. Park's idea of the “marginal man” and his work on cultural and racial hybridity can shed further light on the construction and representation of the hybrid self. The contribution that Park has made to a social theory of hybridity has been overshadowed by research conducted within post-colonial and cultural studies. I do not suggest that recent conceptualisations of hybridity are inadequate; rather that Park has something to contribute to contemporary accounts and in some cases anticipates some of the themes and issues surrounding the concept of hybridity. The following examination connects Park's work on hybridity with ideas such as civilisation, culture and modernity and argues that a mild form of primitivism underlines his notion of the “marginal man”.

Don't tate us! The impediments and drivers of branding museums

This study examines the applicability of brand orientation, and its drivers and impediments in the museum context. A multiple case study approach, with 12 institutions across two countries (United Kingdom and Australia) was carried out. Results suggest that brand orientation provides the cultural platform to retain the distinctiveness of the institution and build the symbolic representation required to remain competitive in the future.

Understanding informations systems outsourcing success and risks through the lens of cognitive biases

Because outsourcing of information systems (IS) is now widespread, it is generally assumed to be successful. It is also often assumed that outsourcing risks are easily managed. In this paper we adopt an “evidence based management” approach to first test these assumptions through a qualitative metaanalysis of academic studies into IS outsourcing outcomes. Our research reveals a shortage of reliable and valid evidence for outsourcing’s benefits, and for the level of risk involved. We then use data from a series of focus groups to explain the paradox of widespread adoption of a strategy with limited empirical support. These focus groups were interpreted through the lens of research on a
range of cognitive mechanisms and biases that are known to affect decision makers. We conclude that cognitive mechanisms that are likely to affect sourcing decisions include framing biases, cognitive dissonance, attribution error, and the “optimism”, “confirmation”, “disconfirmation” and “overconfidence” biases. Given the shortage of supporting evidence, and the potential for these biases to operate, we argue that researchers need to be more critical in their analysis of reports of the success and risks of IS outsourcing.

The performance of solarscan : an automated dermoscopy image analysis instrument for the diagnosis of primary melanoma

Objective To describe the diagnostic performance of SolarScan (Polartechnics Ltd, Sydney, Australia), an automated instrument for the diagnosis of primary melanoma.

Design Images from a data set of 2430 lesions (382 were melanomas; median Breslow thickness, 0.36 mm) were divided into a training set and an independent test set at a ratio of approximately 2:1. A diagnostic algorithm (absolute diagnosis of melanoma vs benign lesion and estimated probability of melanoma) was developed and its performance described on the test set. High-quality clinical and dermoscopy images with a detailed patient history for 78 lesions (13 of which were melanomas) from the test set were given to various clinicians to compare their diagnostic accuracy with that of SolarScan.

Setting Seven specialist referral centers and 2 general practice skin cancer clinics from 3 continents. Comparison between clinician diagnosis and SolarScan diagnosis was by 3 dermoscopy experts, 4 dermatologists, 3 trainee dermatologists, and 3 general practitioners.

Patients Images of the melanocytic lesions were obtained from patients who required either excision or digital monitoring to exclude malignancy.

Main Outcome Measures Sensitivity, specificity, the area under the receiver operator characteristic curve, median probability for the diagnosis of melanoma, a direct comparison of SolarScan with diagnoses performed by humans, and interinstrument and intrainstrument reproducibility.

Results The melanocytic-only diagnostic model was highly reproducible in the test set and gave a sensitivity of 91% (95% confidence interval [CI], 86%-96%) and specificity of 68% (95% CI, 64%-72%) for melanoma. SolarScan had comparable or superior sensitivity and specificity (85% vs 65%) compared with those of experts (90% vs 59%), dermatologists (81% vs 60%), trainees (85% vs 36%; P =.06), and general practitioners (62% vs 63%). The intraclass correlation coefficient of intrainstrument repeatability was 0.86 (95% CI, 0.83-0.88), indicating an excellent repeatability. There was no significant interinstrument variation (P = .80).

Conclusions SolarScan is a robust diagnostic instrument for pigmented or partially pigmented melanocytic lesions of the skin. Preliminary data suggest that its performance is comparable or superior to that of a range of clinician groups. However, these findings should be confirmed in a formal clinical trial.