Introduction to the Synthesis and Purification of Oligonucleotides

Modern nucleic acid synthesizers utilize phosphite triester chemistries that employ stable phosphoramidite monomers to build a growing polymer. These robust reactions allow easy generation of specific oligodeoxyribo‐ and oligoribonucleotides with a variety of labels, modified linkages, and nonstandard bases. Strategies are given for the maximization of synthetic yield, the generation of sequences containing site‐specific modifications, and the isolation of synthetic oligonucleotides. Protocols describe monitoring the progress of synthesis via the trityl assay and methods for deprotection.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/143751/1/cpnca03c.pd

Geometry of bend : singular lines and defects in twist-bend nematics

We describe the geometry of bend distortions in liquid crystals and their fundamental degeneracies, which we call β lines; these represent a new class of linelike topological defect in twist-bend nematics. We present constructions for smecticlike textures containing screw and edge dislocations and also for vortexlike structures of double twist and Skyrmions. We analyze their local geometry and global structure, showing that their intersection with any surface is twice the Skyrmion number. Finally, we demonstrate how arbitrary knots and links can be created and describe them in terms of merons, giving a geometric perspective on the fractionalization of Skyrmions

Design, Synthesis, and Amplification of DNA Pools for In Vitro Selection

Preparation of a random‐sequence DNA pool is presented. The degree of randomization and the length of the random sequence are discussed, as is synthesis of the pool using a DNA synthesizer. Purification of a single‐stranded pool and conversion to a double‐stranded pool are presented as step‐by‐step protocols. Support protocols describe determination of the complexity and skewing of the pool, and optimization of amplification conditions.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/143624/1/cpnc0902.pd

Evaluation of ambr® 250 perfusion bioreactor system as a model for high-throughput perfusion cell culture process development

The efficient development and delivery of high-quality therapeutic products necessitates the need for high-throughput process development (HTPD) tools. In recent years, fed-batch process development timelines have been significantly reduced as industry has implemented fully automated mini bioreactor systems such as the ambr® 250 HT. More recently, as continuous processing for biologics has gained traction, driven by novel technology and economic pressure to significantly improve monoclonal antibody (mAb) production over the standard fed-batch process, more efficient, cost effective, and environmentally sustainable mAb processes are expected. Traditionally, perfusion process development work requires a combination of deep-well plates and small-scale stirred tank bioreactor (STR), both of which are labor intensive and time consuming. The established ambr®250 HT platform has recently been integrated with perfusion capabilities to enable rapid continuous perfusion process development (PD). In this work, we present the results from our assessment of the automated disposable perfusion bioreactor system for high throughput upstream PD activities, including clone selection and process optimization. In addition, several high stress conditions were also examined here to identify optimal operation ranges for the current system. The studies conclude that ambr® 250 perfusion reactor is able to generate process performance and product quality profiles equivalent to bench-top bioreactors for a high cell density perfusion process

Existing climate change will lead to pronounced shifts in the diversity of soil prokaryotes

© The Author(s), 2018. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in mSystems 3 (2018): e00167-18, doi:10.1128/mSystems.00167-18.Soil bacteria are key to ecosystem function and maintenance of soil fertility. Leveraging associations of current geographic distributions of bacteria with historic climate, we predict that soil bacterial diversity will increase across the majority (∼75%) of the Tibetan Plateau and northern North America if bacterial communities equilibrate with existing climatic conditions. This prediction is possible because the current distributions of soil bacteria have stronger correlations with climate from ∼50 years ago than with current climate. This lag is likely associated with the time it takes for soil properties to adjust to changes in climate. The predicted changes are location specific and differ across bacterial taxa, including some bacteria that are predicted to have reductions in their distributions. These findings illuminate the widespread potential of climate change to influence belowground diversity and the importance of considering bacterial communities when assessing climate impacts on terrestrial ecosystems.This work was supported by the Strategic Priority Research Program (XDB15010101, XDA05050404) of the Chinese Academy of Sciences, the National Program on Key Basic Research Project (2014CB954002, 2014CB954004), the National Natural Science Foundation of China (41701298, 41371254), the “135” Plan and Frontiers Projects of Institute of Soil Science (ISSASIP1641), and the National Science and Technology Foundation project (2015FY110100). J.A.G. was supported by the U.S. Dept. of Energy under contract DE-AC02-06CH11357. N.F. was supported by a grant from the National Science Foundation (DEB-0953331). K.S.P. and J.L. were supported by the National Science Foundation (DMS-1069303), the Gordon and Betty Moore Foundation (grant no. 3300), the Gladstone Institutes, and a gift from the San Simeon Fund

Evaluating the impact of Brexit on the pharmaceutical industry.

INTRODUCTION: The UK Pharmaceutical Industry is arguably one of the most important industries to consider in the negotiations following the Brexit vote. Providing tens of thousands of jobs and billions in tax revenue and research investment, the importance of this industry cannot be understated. At stake is the global leadership in the sector, which produces some of the field's most influential basic science and translation work. However, interruptions and losses may occur at multiple levels, affecting patients, researchers, universities, companies and government. GOALS: By understanding the current state of pharmaceutical sector, the potential effect of leaving the European Union (EU) on this successful industry can be better understood. This paper aims to address the priorities for negotiations by collating the analyses of professionals in the field, leading companies and non-EU member states. RESEARCH METHODS: A government healthcare policy advisor and Chief Science Officer (CSO) for a major pharmaceutical firm were consulted to scope the paper. In these discussions, five key areas were identified: contribution, legislative processes, regulatory processes, research and outcomes, commercial risk. Multiple search engines were utilised for selecting relevant material, predominantly PubMed and Google Scholar. To supplement this information, Government documents were located using the "GOV.UK" publications tool, and interviews and commentaries were found through the Google News search function. CONCLUSION: With thorough investigation of the literature, we propose four foundations in the advancement of negotiations. These prioritise: negotiation of 'associated country' status, bilaterally favourable trade agreements, minimal interruption to regulatory bodies and special protection for the movement of workforce in the life sciences industry

Ancestral roles of the Fam20C family of secreted protein kinases revealed in C. elegans.

Fam20C is a secreted protein kinase mutated in Raine syndrome, a human skeletal disorder. In vertebrates, bone and enamel proteins are major Fam20C substrates. However, Fam20 kinases are conserved in invertebrates lacking bone and enamel, suggesting other ancestral functions. We show that FAMK-1, the Caenorhabditis elegans Fam20C orthologue, contributes to fertility, embryogenesis, and development. These functions are not fulfilled when FAMK-1 is retained in the early secretory pathway. During embryogenesis, FAMK-1 maintains intercellular partitions and prevents multinucleation; notably, temperature elevation or lowering cortical stiffness reduces requirement for FAMK-1 in these contexts. FAMK-1 is expressed in multiple adult tissues that undergo repeated mechanical strain, and selective expression in the spermatheca restores fertility. Informatic, biochemical, and functional analysis implicate lectins as FAMK-1 substrates. These findings suggest that FAMK-1 phosphorylation of substrates, including lectins, in the late secretory pathway is important in embryonic and tissue contexts where cells are subjected to mechanical strain

Estimating the global economic benefits of physically active populations over 30 years (2020 to 2050)

OBJECTIVES: We assess the potential benefits of increased physical activity on the global economy for 23 countries and the rest of the world over a 30-year time horizon (from 2020 to 2050). The main factors taken into account in the economic assessment are excess mortality and lower productivity. METHODS: This study links three methodologies. First, we estimate the association between physical inactivity and workplace productivity using multivariable regression models with proprietary data on 120,143 individuals in the UK and six Asian countries (Australia, Malaysia, Hong Kong, Thailand, Singapore and Sri Lanka). Second, we analyse the association between physical activity and mortality risk through a meta-regression analysis with data from 74 prior studies with global coverage. Finally, the estimated effects are combined in a computable general equilibrium (CGE) macroeconomic model to project the economic benefits of physical inactivity over time. RESULTS: Doing at least 150 minutes of moderate-intensity physical activity, as per lower limit of the range recommended by the 2020 World Health Organisation guidelines, would lead to an increase in global GDP of 0.16%-0.23% per year by 2050, worth up to US $314-$446 billion per year and $6.0-$8.6 trillion cumulatively over the 30-year projection horizon (in 2019 prices). The results vary by country due to differences in baseline levels of physical activity and GDP per capita. CONCLUSIONS: Increasing physical activity in the population would lead to reduction in working-age mortality and morbidity and an increase in productivity, particularly through lower presenteeism, leading to substantial economic gains for the global economy

Being Silenced: The Impact of Negative Social Reactions on the Disclosure of Rape

Rape survivors who speak out about their assault experiences are often punished for doing so when they are subjected to negative reactions from support providers. These negative reactions may thereby serve a silencing function, leading some rape survivors to stop talking about their experiences to anyone at all. The current study sought to examine this worst case scenario. Focusing on the qualitative narratives of eight rape survivors who initially disclosed the assault but then stopped disclosing for a significant period of time, this study sought to provide an in-depth description of how negative reactions silenced these survivors. Three routes to silence were identified: 1) negative reactions from professionals led survivors to question whether future disclosures would be effective; 2) negative reactions from friends and family reinforced feelings of self-blame; and 3) negative reactions from either source reinforced uncertainty about whether their experiences qualified as rape. Implications for future research and practice are discussed