ExCyto PCR Amplification

ExCyto PCR cells provide a novel and cost effective means to amplify DNA transformed into competent bacterial cells. ExCyto PCR uses host E. coli with a chromosomally integrated gene encoding a thermostable DNA polymerase to accomplish robust, hot-start PCR amplification of cloned sequences without addition of exogenous enzyme.Because the thermostable DNA polymerase is stably integrated into the bacterial chromosome, ExCyto cells can be transformed with a single plasmid or complex library, and then the expressed thermostable DNA polymerase can be used for PCR amplification. We demonstrate that ExCyto cells can be used to amplify DNA from different templates, plasmids with different copy numbers, and master mixes left on ice for up to two hours. Further, PCR amplification with ExCyto cells is comparable to amplification using commercial DNA polymerases. The ability to transform a bacterial strain and use the endogenously expressed protein for PCR has not previously been demonstrated.ExCyto PCR reduces pipetting and greatly increases throughput for screening EST, genomic, BAC, cDNA, or SNP libraries. This technique is also more economical than traditional PCR and thus broadly useful to scientists who utilize analysis of cloned DNAs in their research

The Emergence of Consensus: a primer

The origin of population-scale coordination has puzzled philosophers and scientists for centuries. Recently, game theory, evolutionary approaches and complex systems science have provided quantitative insights on the mechanisms of social consensus. This paper overviews the main dimensions over which the debate has unfolded and discusses some representative results, with a focus on those situations in which consensus emerges `spontaneously' in absence of centralised institutions. Covered topics include the macroscopic consequences of the different microscopic rules of behavioural contagion, the role of social networks, and the mechanisms that prevent the formation of a consensus or alter it after it has emerged. Special attention is devoted to the recent wave of experiments on the emergence of consensus in social systems

"Meaning" as a sociological concept: A review of the modeling, mapping, and simulation of the communication of knowledge and meaning

The development of discursive knowledge presumes the communication of meaning as analytically different from the communication of information. Knowledge can then be considered as a meaning which makes a difference. Whereas the communication of information is studied in the information sciences and scientometrics, the communication of meaning has been central to Luhmann's attempts to make the theory of autopoiesis relevant for sociology. Analytical techniques such as semantic maps and the simulation of anticipatory systems enable us to operationalize the distinctions which Luhmann proposed as relevant to the elaboration of Husserl's "horizons of meaning" in empirical research: interactions among communications, the organization of meaning in instantiations, and the self-organization of interhuman communication in terms of symbolically generalized media such as truth, love, and power. Horizons of meaning, however, remain uncertain orders of expectations, and one should caution against reification from the meta-biological perspective of systems theory

Structural Variants Create New Topological-Associated Domains and Ectopic Retinal Enhancer-Gene Contact in Dominant Retinitis Pigmentosa

The cause of autosomal-dominant retinitis pigmentosa (adRP), which leads to loss of vision and blindness, was investigated in families lacking a molecular diagnosis. A refined locus for adRP on Chr17q22 (RP17) was delineated through genotyping and genome sequencing, leading to the identification of structural variants (SVs) that segregate with disease. Eight different complex SVs were characterized in 22 adRP-affected families with >300 affected individuals. All RP17 SVs had breakpoints within a genomic region spanning YPEL2 to LINC01476. To investigate the mechanism of disease, we reprogrammed fibroblasts from affected individuals and controls into induced pluripotent stem cells (iPSCs) and differentiated them into photoreceptor precursor cells (PPCs) or retinal organoids (ROs). Hi-C was performed on ROs, and differential expression of regional genes and a retinal enhancer RNA at this locus was assessed by qPCR. The epigenetic landscape of the region, and Hi-C RO data, showed that YPEL2 sits within its own topologically associating domain (TAD), rich in enhancers with binding sites for retinal transcription factors. The Hi-C map of RP17 ROs revealed creation of a neo-TAD with ectopic contacts between GDPD1 and retinal enhancers, and modeling of all RP17 SVs was consistent with neo-TADs leading to ectopic retinal-specific enhancer-GDPD1 accessibility. qPCR confirmed increased expression of GDPD1 and increased expression of the retinal enhancer that enters the neo-TAD. Altered TAD structure resulting in increased retinal expression of GDPD1 is the likely convergent mechanism of disease, consistent with a dominant gain of function. Our study highlights the importance of SVs as a genomic mechanism in unsolved Mendelian diseases

Indigenous biosecurity: Māori responses to kauri dieback and myrtle rust in Aotearoa New Zealand

It is widely acknowledged that Indigenous peoples have traditional knowledge relevant to modern environmental management. By asserting roles within associated science and policy networks, such Indigenous Knowledge (IK) can be seen as part of the resistance to colonisation that includes protest, treaty making, political and economic empowerment, legislation, cultural renaissance and regulatory influence. In New Zealand, these achievements inform attempts by Māori (the Indigenous people of New Zealand) to manage forest ecosystems and cultural keystone species. This chapter presents two case studies of how indigenous participation in modern biosecurity through the example of Māori asserting and contributing to forest management. While progress is often frustratingly slow for indigenous participants, significant gains in acceptance of Māori cultural frameworks have been achieved

Embryonic Diapause Is Conserved across Mammals

Embryonic diapause (ED) is a temporary arrest of embryo development and is characterized by delayed implantation in the uterus. ED occurs in blastocysts of less than 2% of mammalian species, including the mouse (Mus musculus). If ED were an evolutionarily conserved phenomenon, then it should be inducible in blastocysts of normally non-diapausing mammals, such as domestic species. To prove this hypothesis, we examined whether blastocysts from domestic sheep (Ovis aries) could enter into diapause following their transfer into mouse uteri in which diapause conditions were induced. Sheep blastocysts entered into diapause, as demonstrated by growth arrest, viability maintenance and their ED-specific pattern of gene expression. Seven days after transfer, diapausing ovine blastocysts were able to resume growth in vitro and, after transfer to surrogate ewe recipients, to develop into normal lambs. The finding that non-diapausing ovine embryos can enter into diapause implies that this phenomenon is phylogenetically conserved and not secondarily acquired by embryos of diapausing species. Our study questions the current model of independent evolution of ED in different mammalian orders

The effectiveness of physical activity monitoring and distance counselling in an occupational health setting - a research protocol for a randomised controlled trial (CoAct)

<p>Abstract</p> <p>Background</p> <p>The CoAct (Cocreating Activity) study is investigating a novel lifestyle intervention, aimed at the working population, with daily activity monitoring and distance counselling via telephone and secure web messages. The main purpose of this study is to evaluate the effectiveness of lifestyle counselling on the level of physical activity in an occupational health setting. The purposes include also analysing the potential effects of changes in physical activity on productivity at work and sickness absence, and healthcare costs. This article describes the design of the study and the participant flow until and including randomization.</p> <p>Methods/Design</p> <p>CoAct is a randomised controlled trial with two arms: a control group and intervention group with daily activity monitoring and distance counselling. The intervention focuses on lifestyle modification and takes 12 months. The study population consists of volunteers from 1100 eligible employees of a Finnish insurance company. The primary outcomes of this study are change in physical activity measured in MET minutes per week, work productivity and sickness absence, and healthcare utilisation. Secondary outcomes include various physiological measures. Cost-effectiveness analysis will also be performed. The outcomes will be measured by questionnaires at baseline, after 6, 12, and 24 months, and sickness absence will be obtained from the employer's registers.</p> <p>Discussion</p> <p>No trials are yet available that have evaluated the effectiveness of daily physical activity monitoring and distance counselling in an occupational health setting over a 12 month period and no data on cost-effectiveness of such intervention are available.</p> <p>Trial Registration</p> <p>ClinicalTrials.gov identifier: NCT00994565</p

Antimicrobial Properties of Ethylene Vinyl Alcohol/Epsilon-Polylysine Films and Their Application in Surimi Preservation

[EN] Polymer films based on ethylene vinyl copolymers (EVOH) containing a 29 % (EVOH 29) and a 44 % molar percentage of ethylene (EVOH 44), and incorporating epsilon-polylysine (EPL) at 0 %, 1 %, 5 % and 10 % were successfully made by casting. The optical properties and the amount of EPL released from the films to phosphate buffer at pH 7.5 were evaluated, films showing great transparency and those of EVOH 29 copolymer releasing a greater amount of EPL. The antimicrobial properties of the resulting films were tested in vitro against different foodborne microorganisms and in vivo in surimi sticks. With regard to the antimicrobial capacity tested in vitro in liquid medium at 37 A degrees C and 4 A degrees C against Listeria monocytogenes and Escherichia coli over a period of 72 h, films showed a considerable growth inhibitory effect against both pathogens, more notably against L. monocytogenes, and being EVOH 29 more effective than EVOH 44 films. At 37 A degrees C, total growth inhibition was observed for EVOH 29 films incorporating 10 % EPL against both microorganisms whereas the copolymer EVOH 44 did show total inhibition against L. monocytogenes and the growth of E. coli was reduced by 6.64 log units. At 4 A degrees C, no film was able to inhibit completely bacterial growth. Scanning electron microscopy micrographs showed corrugated cell surfaces with blisters and bubbles, and collapse of the cells appearing shorter and more compact after treatment with EPL. Finally, the films were successfully used to increase the shelf life of surimi sticks. The results show the films developed have a great potential for active food packaging applications.The authors acknowledge the financial support of the Spanish Ministry of Economy and Competitiveness, projects AGL2012-39920-C03-01, and fellowship funding for V. M.-G.Muriel-Galet, V.; Lopez-Carballo, G.; Gavara Clemente, R.; Hernández-Muñoz, P. (2014). 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Marine Biodiversity in South Africa: An Evaluation of Current States of Knowledge

Continental South Africa has a coastline of some 3,650 km and an Exclusive Economic Zone (EEZ) of just over 1 million km2. Waters in the EEZ extend to a depth of 5,700 m, with more than 65% deeper than 2,000 m. Despite its status as a developing nation, South Africa has a relatively strong history of marine taxonomic research and maintains comprehensive and well-curated museum collections totaling over 291,000 records. Over 3 million locality records from more than 23,000 species have been lodged in the regional AfrOBIS (African Ocean Biogeographic Information System) data center (which stores data from a wider African region). A large number of regional guides to the marine fauna and flora are also available and are listed

A Model of Salmonella Colitis with Features of Diarrhea in SLC11A1 Wild-Type Mice

Background: Mice do not get diarrhea when orally infected with S. enterica, but pre-treatment with oral aminoglycosides makes them susceptible to Salmonella colitis. However, genetically susceptible ItyS mice (Nramp1 G169D allele) die from systemic infection before they develop diarrhea, so a new model is needed to study the pathogenesis of diarrhea. We pretreated ItyR mice (Nramp1 G169) with oral kanamycin prior to infecting them with virulent S. Typhimurium strain 14028s in order to study Salmonella-induced diarrhea. We used both a visual scoring system and the measurement of fecal water content to measure diarrhea. BALB/c.D2 Nramp1 congenic started losing weight 5 days post-infection and they began to die from colitis 10–14 days after infection. A SPI-1 (invA) mutant caused cecal, but not colonic inflammation and did not cause diarrhea. A phoP- mutant did not cause manifestations of diarrhea in either normal or NADPHdeficient (gp91 phox) mice. However, strain 14028s caused severe colitis and diarrhea in gp91 phox-deficient mice on an ItyR background. pmr A and F mutants, which are less virulent in orally infected BALB/c mice, were fully virulent in this model of colitis. Conclusions: S. enterica must be able to invade the colonic epithelium and to persist in the colon in order to cause colitis with manifestations of diarrhea. The NADPH oxidase is not required for diarrhea in Salmonella colitis. Furthermore,