Polymerase II Promoter Strength Determines Efficacy of microRNA Adapted shRNAs

Since the discovery of RNAi and microRNAs more than 10 years ago, much research has focused on the development of systems that usurp microRNA pathways to downregulate gene expression in mammalian cells. One of these systems makes use of endogenous microRNA pri-cursors that are expressed from polymerase II promoters where the mature microRNA sequence is replaced by gene specific duplexes that guide RNAi (shRNA-miRs). Although shRNA-miRs are effective in directing target mRNA knockdown and hence reducing protein expression in many cell types, variability of RNAi efficacy in cell lines has been an issue. Here we show that the choice of the polymerase II promoter used to drive shRNA expression is of critical importance to allow effective mRNA target knockdown. We tested the abundance of shRNA-miRs expressed from five different polymerase II promoters in 6 human cell lines and measured their ability to drive target knockdown. We observed a clear positive correlation between promoter strength, siRNA expression levels, and protein target knockdown. Differences in RNAi from the shRNA-miRs expressed from the various promoters were particularly pronounced in immune cells. Our findings have direct implications for the design of shRNA-directed RNAi experiments and the preferred RNAi system to use for each cell type

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Correction to: Cluster identification, selection, and description in Cluster randomized crossover trials: the PREP-IT trials

An amendment to this paper has been published and can be accessed via the original article

Advancing Racial Equity in Philanthropy: A Scan of Philanthropy-Serving Organizations

United Philanthropy Forum conducted a scan of regional and national philanthropy-serving organizations (PSOs) in February through May 2018 to get a more comprehensive understanding of PSOs' current work and future needs to advance racial equity in philanthropy. The scan involved both a survey that asked about PSOs' current work, future needs and greatest challenges in advancing racial equity, plus in-depth interviews to discuss what it takes to do this work effectively and to identify their key challenges, barriers and opportunities for addressing systemic inequities.The scan reflects the input of 43 regional and national PSOs that participated in the scan survey and/or the scan interviews, representing more than half of the Forum's membership. The scan report includes the Forum's plan for action to respond to the scan's key findings.The Forum's racial equity scan was made possible in part thanks to support from the Ford Foundation and W.K. Kellogg Foundation

Is Infant/Toddler Anemia a Problem across Rural China? A Mixed-Methods Analysis

In the past, iron-deficiency anemia in children has had a widespread presence in rural China. Given the recent economic growth in China, it is unclear if anemia among infants/toddlers remains a problem. The objective of this study is to measure the anemia rate in rural Chinese infants/toddlers across four major subpopulations and attempt to discover the sources of anemia. We use a mixed-methods approach combining quantitative data on 2909 rural Chinese infants/toddlers and their families with qualitative interviews with 84 caregivers of infants aged 6 to 30 months. Quantitative analysis indicates that the overall prevalence of anemia (43%) within sampled infants/toddlers was high, especially in comparison to the low rates of stunting (2–5%), being underweight (2%), and wasting (2–4%). These findings suggest that in rural China, anemia stems from the poor quality of the diets of infants/toddlers, rather than insufficient quantities of food being consumed. Qualitative analysis illustrates the factors that are contributing to anemia. Caregivers do not understand the causes of this condition, the symptoms that would lead one to recognize this condition, or the steps needed to treat their child with this condition. The findings offer a comprehensive understanding of the limited awareness of anemia among rural Chinese caregivers

Implementing stakeholder engagement to explore alternative models of consent: An example from the PREP-IT trials

Introduction: Cluster randomized crossover trials are often faced with a dilemma when selecting an optimal model of consent, as the traditional model of obtaining informed consent from participant's before initiating any trial related activities may not be suitable. We describe our experience of engaging patient advisors to identify an optimal model of consent for the PREP-IT trials. This paper also examines surrogate measures of success for the selected model of consent. Methods: The PREP-IT program consists of two multi-center cluster randomized crossover trials that engaged patient advisors to determine an optimal model of consent. Patient advisors and stakeholders met regularly and reached consensus on decisions related to the trial design including the model for consent. Patient advisors provided valuable insight on how key decisions on trial design and conduct would be received by participants and the impact these decisions will have. Results: Patient advisors, together with stakeholders, reviewed the pros and cons and the requirements for the traditional model of consent, deferred consent, and waiver of consent. Collectively, they agreed upon a deferred consent model, in which patients may be approached for consent after their fracture surgery and prior to data collection. The consent rate in PREP-IT is 80.7%, and 0.67% of participants have withdrawn consent for participation. Discussion: Involvement of patient advisors in the development of an optimal model of consent has been successful. Engagement of patient advisors is recommended for other large trials where the traditional model of consent may not be optimal