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

BackgroundThe 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.ResultsTo 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.ConclusionWe 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.<br /

Prevention of benzene-induced myelotoxicity by nonsteroidal anti-inflammatory drugs.

Benzene affects hematopoietic progenitor cells leading to bone marrow depression and genotoxic effects such as micronucleus formation. Progenitor cell proliferation and differentiation are inhibited by prostaglandins produced by macrophages. Administration of benzene to DBA/2 or C57BL/6 mice caused a dose-dependent bone marrow depression and a significant increase in marrow prostaglandin E level and both were prevented by the coadministration of indomethacin and other inhibitors of the cyclooxygenase component of prostaglandin H synthase. Levels of benzene that decreased bone marrow cellularity also caused genotoxic effects measured as increased micronucleated polychromatic erythrocytes in peripheral blood, which was also prevented by the coadministration of indomethacin. These results suggest a possible role for prostaglandin synthase in benzene myelotoxicity; a mechanism by which this might occur is presented

Hard loss of stability in Painlev\'e-2 equation

A special asymptotic solution of the Painlev\'e-2 equation with small parameter is studied. This solution has a critical point $t_*$ corresponding to a bifurcation phenomenon. When $t<t_*$ the constructed solution varies slowly and when $t>t_*$ the solution oscillates very fast. We investigate the transitional layer in detail and obtain a smooth asymptotic solution, using a sequence of scaling and matching procedures

Homogeneity of antimicrobial policy, yet heterogeneity of antimicrobial resistance: antimicrobial non-susceptibility among 108,717 clinical isolates from primary, secondary and tertiary care patients in London

Objectives: We examined the four-year trend in antimicrobial susceptibilities and prescribing across levels-of-care at two London teaching hospitals and their multisite renal unit, and for the surrounding community. Methods: Laboratory and pharmacy information management systems were interrogated, with antimicrobial use and susceptibilities analysed between hospitals, within hospitals, and over time. Results: 108,717 isolates from 71,687 patients were identified, with significant differences (at p<0.05) in antimicrobial susceptibility between and within hospitals. Across the four years, rates of extended-spectrum β-lactamase (ESBL)-/AmpC-producing Enterobacteriaceae ranged from 6.4 to 10.7% among community isolates, 17.8 to 26.9% at ward level and 25.2 to 52.5% in critical-care. Significant variations were also demonstrated in glycopeptide-resistant enterococci (ward level 6.2 to 17.4%; critical-care 21.9 to 56.3%), methicillin-resistant Staphylococcus aureus (ward level 18.5 to 38.2%; critical-care 12.5 to 47.9%) and carbapenem-resistant Pseudomonas spp. (ward level 8.3 to 16.9%; critical-care 19.9 to 53.7%). Few instances of persistently higher resistance were seen between the hospitals in equivalent cohorts, despite persistently higher antimicrobial use in hospital 1 than hospital 2. We found significant fluctuations in non-susceptibility year-on-year across the cohorts, but with few persistent trends. Conclusions: The marked heterogeneity of antimicrobial susceptibilities between hospitals, within hospitals, and over time demands detailed, standardised surveillance and appropriate benchmarking to identify possible drivers and effective interventions. Homogenous antimicrobial policies are unlikely to continue to be suitable as individual hospitals join hospital networks, and policies should be tailored to local resistance rates, at least at the hospital level, and possibly with finer resolution, particularly for critical-care

The politics of the teaching of reading

Historically, political debates have broken out over how to teach reading in primary schools and infant classrooms. These debates and “reading wars” have often resulted from public concerns and media reportage of a fall in reading standards. They also reflect the importance placed on learning to read by parents, teachers, employers, and politicians. Public and media-driven controversies over the teaching of reading have resulted in intense public and professional debates over which specific methods and materials to use with beginning readers and with children who have reading difficulties. Recently, such debates have led to a renewed emphasis on reading proficiency and “standardized” approaches to teaching reading and engaging with literacy. The universal acceptance of the importance of learning to read has also led to vested interests in specific methods, reading programmes, and early literacy assessments amongst professional, business, commercial, and parental lobbying groups. This article traces these debates and the resulting growing support for a quantitative reductionist approach to early-reading programmes

The luminous type Ia supernova 2022ilv and its early excess emission

We present observations and analysis of the host-less and luminous type Ia supernova 2022ilv, illustrating it is part of the 2003fg-like family, often referred to as super-Chandrasekhar (Ia-SC) explosions. The ATLAS light curve shows evidence of a short-lived, pulse-like early excess, similar to that detected in another luminous type Ia supernova (SN 2020hvf). The light curve is broad and the early spectra are remarkably similar to SN 2009dc. Adopting a redshift of $z=0.026 \pm 0.005$ for SN 2022ilv based on spectral matching, our model light curve requires a large $^{56}$Ni mass in the range $0.7-1.5$ M$_{\odot}$, and a large ejecta mass in the range $1.6-2.3$ M$_{\odot}$. The early excess can be explained by fast-moving SN ejecta interacting with a thin, dense shell of circumstellar material close to the progenitor ($\sim 10^{13}$ cm), a few hours after the explosion. This may be realised in a double-degenerate scenario, wherein a white dwarf merger is preceded by ejection of a small amount ($\sim 10^{-3}-10^{-2}$ M$_{\odot}$) of hydrogen and helium-poor tidally stripped material. A deep pre-explosion Pan-STARRS1 stack indicates no host galaxy to a limiting magnitude of $r \sim 24.5$. This implies a surprisingly faint limit for any host of $M_r \gtrsim -11$, providing further evidence that these types of explosion occur predominantly in low-metallicity environments.Comment: Accepted to ApJL after minor revisio

Processfolio: uniting Academic Literacies and Critical Emancipatory Action Research for practitioner-led inquiry into EAP writing assessment

This paper reports on the design and implementation of an alternative form of writing assessment on a UK English for Academic Purposes (EAP) presessional course. The assessment, termed processfolio, was a response to research inquiry into how writing assessment in a local context negated student agency and inculcated disempowering models of teaching and learning academic writing. The project merged an Academic Literacies approach to writing (Lea and Street, 1998) with a Critical Emancipatory Action Research (Carr and Kemmis, 1986) framework and a Critical Realist(Bhaskar, 1989) perspective. Data collected from the folios and interviews with students and teachers on their experiences of the processfolio found that a small scale intervention has potential for agency to be exercised within the highly constrained context of a UK EAP pre-sessional. New directions in research are proposed which can engage students and teachers to work for change in UK EAP assessment within their internal and external constraints

Marking gender studies:the (Radical) value of creative-critical assessment

Feminist pedagogies have established the need to query power structures in terms of curriculum content and teaching praxis. However, the topic of student assessment poses difficulties: it is a means through which students’ performance is evaluated and quantified according to set institutionalised criteria that values particular forms of hegemonic knowledge. The following article presents a self-reflexive exploration of assessment within a Gender Studies module taught in the Autumn semesters of the 2017/18 and 2018/19 academic years at a UK university. The module was a core component of the institution’s MA in Gender Studies. This was an exciting opportunity to experiment with assessment styles corresponding to feminist pedagogies to help develop students’ and instructors’ disciplinary scope and explore the radical potential for creative-critical approaches to assessment. This article outlines some the challenges of employing alternative modes of learning and teaching from a feminist perspective and suggests some strategies to address these

The Discovery Potential of a Super B Factory

The Proceedings of the 2003 SLAC Workshops on flavor physics with a high luminosity asymmetric e+e- collider. The sensitivity of flavor physics to physics beyond the Standard Model is addressed in detail, in the context of the improvement of experimental measurements and theoretical calculations.Comment: 476 pages. Printed copies may be obtained by request to [email protected] . arXiv admin note: v2 appears to be identical to v

The Surgical Infection Society revised guidelines on the management of intra-abdominal infection

Background: Previous evidence-based guidelines on the management of intra-abdominal infection (IAI) were published by the Surgical Infection Society (SIS) in 1992, 2002, and 2010. At the time the most recent guideline was released, the plan was to update the guideline every five years to ensure the timeliness and appropriateness of the recommendations. Methods: Based on the previous guidelines, the task force outlined a number of topics related to the treatment of patients with IAI and then developed key questions on these various topics. All questions were approached using general and specific literature searches, focusing on articles and other information published since 2008. These publications and additional materials published before 2008 were reviewed by the task force as a whole or by individual subgroups as to relevance to individual questions. Recommendations were developed by a process of iterative consensus, with all task force members voting to accept or reject each recommendation. Grading was based on the GRADE (Grades of Recommendation Assessment, Development, and Evaluation) system; the quality of the evidence was graded as high, moderate, or weak, and the strength of the recommendation was graded as strong or weak. Review of the document was performed by members of the SIS who were not on the task force. After responses were made to all critiques, the document was approved as an official guideline of the SIS by the Executive Council. Results: This guideline summarizes the current recommendations developed by the task force on the treatment of patients who have IAI. Evidence-based recommendations have been made regarding risk assessment in individual patients; source control; the timing, selection, and duration of antimicrobial therapy; and suggested approaches to patients who fail initial therapy. Additional recommendations related to the treatment of pediatric patients with IAI have been included. Summary: The current recommendations of the SIS regarding the treatment of patients with IAI are provided in this guideline