bluuebunny/crossref_metadata_2025 huggingface dataset:
Seasonal variation in thoracic vessel calcifications: Evidence from a chest computed tomography study
Authors: Vehmas Tapio , Hiltunen Asta , Leino-Arjas Päivi
Published: January 2010

Background: Cardiovascular disease incidence and mortality exhibit a winter peak and a summer trough, a fact that could have radiological manifestations.

Purpose: To identify possible seasonal trends in the occurrence of thoracic vessel calcifications.

Material and Methods: 505 male construction workers (aged 39–80 years) were each imaged once with chest spiral computed tomography (CT) during a 2-year period. Based on visual assessment of calcified plaques (0=no, 1=slight, 2=moderate, 3=extensive calcification), sum scores of atherosclerosis in coronary arteries, in the thoracic aorta, in the pre-cervical artery bases, and overall were constructed. The scores were regressed on the annual rank number of the CT day.

Results: By using the cubic regression model, seasonal variation in calcified plaques in coronary arteries ( P=0.003), in pre-cervical artery origins ( P=0.015), and in the overall sum score ( P=0.004) was observed. The peak occurred in January–February and the nadir in August. Depending on the model, about 2–3% of the variation in atherosclerotic calcifications could be explained by the season of imaging.

Conclusion: The observed seasonal trend in calcifications parallels with mortality reports. Seasonal variations should be considered in atherosclerosis treatment studies. Confirmatory studies using modern imaging technology are needed in different countries and geographical locations, preferably with repeat imaging of the same individuals.
Age coding: On age-based practices of distinction
Authors: Krekula Clary
Published: January 2010

In this article I discuss how conceptions of age create individuals’ subjective experiences of old age. I introduce the concept of age coding and argue that it should be understood as referring to practices of distinction that are based on and preserve representations of actions, phenomena, and characteristics as associated with and applicable to demarcated ages. The article illustrates how age codes can be used (1) as age norms; (2) to legitimize, negotiate and regulate symbolic and material resources; (3) as a resource in interactions; as well as (4) to create age-based norms and deviance. This broad application means that the concept should be understood as a tool for analyzing age relations generally.
Objectifying User Critique
Authors: Mahler V. , Prokosch H. U. , Oschem M.
Published: January 2010

Objectives: The aim of this study is to objectify user critique rendering it usable for quality assurance. Based on formative and summative evaluation results we strive to promote software improvements; in our case, the physician discharge letter composition process at the Department of Dermatology, University Hospital Erlangen, Germany.

Methods: We developed a novel six-step approach to objectify user critique: 1) acquisition of user critique using subjectivist methods, 2) creation of a workflow model, 3) definition of hypothesis and indicators, 4) measuring of indicators, 5) analyzing results, 6) optimization of the system regarding both subjectivist and objectivist evaluation results. In particular, we derived indicators and workflows directly from user critique/narratives. The identified indicators were mapped onto work-flow activities, creating a link between user critique and the evaluated system.

Results: Users criticized a new discharge letter system as “too slow” and “too labor-intensive” in comparison with the previously used system. In a stepwise approach we collected subjective user critique, derived a comprehensive process model including deviations and deduced a set of five indicators for objectivist evaluation: processing time, system-related waiting time, number of mouse clicks, number of keyboard inputs, and throughput time. About 3500 measurements have been performed to compare the work-flow-steps of both systems, regarding 20 discharge letters.

Although the difference of the mean total processing time between both systems was statistically insignificant (2011.7 s vs. 1971.5 s; p = 0.457), we detected a significant difference in waiting times (101.8 s vs. 37.2 s; p < 0.001) and number of user interactions (77 vs. 69; p < 0.001) in favor of the old system, thus objectifying user critique.

Conclusions: Our six-step approach enables objectification of user critique, resulting in objective values for continuous quality assurance. To our knowledge no previous study in medical informatics mapped user critique onto workflow steps. Subjectivist analysis prompted us to use the indicator system-related waiting time for the objectivist study, which was rarely done before. We consider combining subjectivist and objectivist methods as a key point of our approach. Future work will concentrate on automated measurement of indicators.
Balma Guilanyà (Prepirineo de Lleida) y el Aziliense en el noreste de la Península Ibérica
Authors: Martínez-Moreno Jorge , Mora Torcal Rafael
Published: January 2010

Guilanyà es un asentamiento localizado en las primeras estribaciones del Prepirineo de Lleida, a 1.150 m de altitud. En este artículo se exponen las observaciones preliminares derivadas de un sondeo realizado en el verano del 2008, que ha permitido definir los niveles arqueológicos K y EJ. Paralelamente, se presentan nuevas dataciones por 14C AMS que contextualizan cronométricamente la secuencia Tardiglaciar de este yacimiento. Estos resultados permiten identificar la gestión recurrente de los ecosistemas de montaña durante la mejora Bolling/Allerod. Las características tecno-tipológicas que se reconocen en estos niveles nos permiten asignarlos al Aziliense, tradición cultural escasamente visualizada en la vertiente sur de los Pirineos.
La Pedra da Póvoa (Trás-os-Montes, Portugal). Una pieza escultórica de la Edad del Hierro
Authors: Fonte João , Santos Estévez Manuel , Bacelar Alves Lara , López Noia Raquel
Published: January 2010

Se da a conocer la Pedra da Póvoa (Póvoa, Ribeira de Pena), un monolito inédito, grabado con motivos abstractos y un zoomorfo. Consideramos posible fecharlo en la Edad del Hierro del Noroeste Peninsular. Se presentan las condiciones del hallazgo, emplazamiento y descripción tecno-morfológica de los grabados y se plantean algunas consideraciones en torno a su contextualización crono-estilística.
Ámbitos sociales compartidos: convergencia evolutiva en la Prehistoria y la práctica contemporánea
Authors: Kohl Philip L.
Published: January 2010

En este artículo distingo entre perspectivas evolucionistas e históricas sobre el pasado, adoptando el concepto de “ámbitos sociales” para defender una interpretación macrohistórica del registro arqueológico. La unidad de análisis no es una cultura o civilización arqueológica sino los grupos sociales implicados de manera inextricable con otros grupos en interconexiones reticulares en las que las tecnologías son difundidas y modificadas por otros grupos atrapados en esos mismos procesos. Tales interconexiones pueden ser trazadas mejor arqueológicamente examinando la expansión de las tecnologías y las prácticas subsistenciales. Otras perspectivas macrohistóricas sobre el pasado, como el análisis de los sistemas mundiales, a menudo exigen demasiado del registro arqueológico y son empleadas de manera anacrónica. La Prehistoria documenta la siempre creciente participación de los grupos en ámbitos sociales que, finalmente, convergen. Concluyo haciendo hincapié en la necesidad de una perspectiva acerca del pasado que resalte su naturaleza compartida en la que todos los pueblos han contribuido a las interacciones con sus vecinos y se han beneficiado de ellas.
SOME NEW SPECIES OF TYPHLOCYBA (HOMOPTERA, CICADELLIDAE)
Authors: Beamer R. H.
Published: January 2010

Resembling T. gillettei but somewhat larger. Two inner pairs of processes of aedeagus of about same length with innner pair not bent at middle and dorsal spine as long as shaft is wide. Length 4.5 mm.

Color: As in heavily marked T. gillettei V. D.
A REMARKABLE INSTANCE OF RETENTION OF A DOUBLE SPERMATHECA IN A DOLICHOPSYLLID FLEA, OPISOCROSTIS BRUNERI (BAKER).
Authors: Holland George P.
Published: January 2010

In some species of insects there is only a single mating, but the egg-laying period may extend over a considerable length of time. Under such circumstances there must be some provision in the female for the storage and preservation of the supply of seminal fluid received during coitus, so that spermatozoa will be available as eggs mature. This is generally effected by an organ known as the spermatheca or receptaculum seminis, a flask-like structure which may open by means of a duct directly into the vagina, but more frequently empties into a bursa copulatrix, which in turn opens into the vagina.
NOTES ON SOME FLEAS OF NEW YORK STATE
Authors: Jameson E. W.
Published: January 2010

During the course of field work with small mammals in central New York, the writer made a collection of fleas. Those records which give additional information on the host distribution of these parasites, are listed below. Some significant records from the Cornell University collection are also cited. This short list will serve to supplement the checklist of fleas for New York State of Stewart (1933, Jour. N. Y. Ent. Soc., 41:253-262). The availability of the Cornell collection was made through the kindness of Dr. Robert Matheson, and to him I tender my thanks.
ON THE NEARCTIC SPECIES OF MOTES (HYMENOPTERA, SPHECIDAE)
Authors: Pate V. S. L.
Published: January 2010

The dominant genus of Larrine wasps in warm countries throughout the world is Motes Kohl (olim Notogonia Costa seu Notogonidea Rohwer), but it is rather poorly represented in the Nearctic fauna. Hitherto only five species have been recorded from the United States: argentatus (Palisot de Beauvois), 1811, aequalis (Fox), 1893, nigripennis (Fox), 1893, occidentalis (Viereck), 1902, and subaequalis (Rohwer) 1909. To this roster may now be added the following two very distinctive southern forms.
NOTES ON THE BETULELLA GROUP OF DEPRESSARIA, WITH DESCRIPTION OF NEW SPECIES. (LEPIDOPTERA, OECOPHORIDAE)
Authors: McDunnough J.
Published: January 2010

In his recent Revision of the Oecophoridae (1941, Proc. U. S. N. M. Vol. 90, No. 3107) Gates Clarke (p. 196) records the larva of Depressaria betulella Bsk. as feeding on both Betula and Ostrya. As material bred by us from Ostrya this past summer in the Ottawa region did not at all match with our series of the birch-feeder I have made a genitalic study of the species of this group in the Canadian National Collection in order to check up on the matter.
THREE ASTER-FEEDING COLEOPHORIDS AND THEIR ALLIES (LEPIDOPTERA)
Authors: McDunnough J.
Published: January 2010

In the past two years three distinct types of Coleophorid cases have been found in the Ottawa region in the early spring, attached mostly to the leaves of Aster tradescanthi or allied, thin-leafed species. Small series of moths have been bred from all three types of cases, the adults appearing in late June or early July.
NEW MICROLEPIDOPTERA WITH NOTES
Authors: McDunnough J.
Published: January 2010

For some time it has appeared to me that two species have been going under the above name. Heinrich in his revision gives the food-plant as Hamamelis but the few bred specimens in our collection from southern Ontario from larvae on this plant together with several captured specimens from New York State did not match Kearfott's description particularly well.
CAMPOPLEGINE NOTES AND DESCRIPTIONS (HYMENOPTERA, ICHNEUMONIDAE)
Authors: Walley G. Stuart
Published: January 2010

The following notes deal with three species in the genera Spudastica, Echthronomas and Campoctonus, and are based on material in the National Collection, supplemented by specimens loaned from the United States National Museum and the personal collection of Dr. H. K. Townes, to whom I am especially indebted for providing me with this additional material.
TWO NEW RACES OF THECLINAE FROM SOUTHERN BRITISH COLUMBIA (LEPIDOPTERA)
Authors: McDunnough J.
Published: January 2010

Differs from typical behrii (as represented by specimens from Mammoth Camp, Mono Co., Calif., and by Holland's figure) in the considerably larger size, the much darker color of the underside and the heavy nature of the black spotting. On both primaries and secondaries the postdiscal row of spots is much more pronounced and on secondaries the submarginal row is very heavy and the spots show a distinct tendency to inward, triangular elongation.
AN INTERESTING OCCURRENCE OF MUSCA DOMESTICA L. LARVAE IN INFANT BEDDING
Authors: Chapman R. Keith
Published: January 2010

On August 10, 1943, the Guelph sanitary inspector brought into the Entomological Laboratory at the Ontario Agricultural College an envelope containing six dipterous maggots taken from a baby's mattress. The mother of the infant was very upset, as she had no idea what the “things” crawling out of her baby's mattress were, and was considering taking some action against the furniture company from which she had purchased the mattress only ten days previously.
THE IDENTITY OF A BORER ATTACKING PEACH TREES IN THE OKANAOAN VALLEY OF BRITISH COLUMBIA
Authors: Venables E. P.
Published: January 2010

Peach trees throughout the Okanagan are frequently attacked by a borer which has been generally referred to in the past as Sanninoidea exitiosa Say. It is doubtful, however, if specimens of it were ever submitted for authoritative determination prior to 1940.
CONTRIBUTIONS TO A KNOWLEDGE OF THE LARVAL ELATERIDAE (COLEOPTERA): NO. 3; AGRIOTES ESCH. AND DALOPIUS ESCH.
Authors: Glen Robert
Published: January 2010

As our knowledge of the larvae increases, the fundamental differences between Agriotes Eschscholtz and Dalopius Eschscholtz become more apparent. However, the larval characters hitherto used to separate the genera are now known to be quite inadequate.
A NOTE ON DDT, THE NEW SYNTHETIC INSECTICIDE
Authors: Twinn C. R.
Published: January 2010

The following statement concerning dichloro-diphenyl-trichlorethane, or 2, 2-bis (parachlorophenyl)-1,1,1-trichloroethane, more conveniently referred to as DDT, has been prepared in view of the widespread interest among entomologists in this product and in its pest control possibilities. It is based on information obtained from various sources.
NEW NEARCTIC CRANE-FLIES (TIPULIDAE, DIPTERA). PART XXV
Authors: Alexander Charles P.
Published: January 2010

The preceding instalment under this title was published in 1945 (Can. Ent., 77: 140-144). At this time I am including the description of five species of the genus Tipula from Washington, Oregon and California, collected by Messrs. T. H. G. Aitken, Mont A. Cazier, J. A. Macnab and A. L. Melander, to whom I am greatly indebted for the privilege of studying very extensive series of Tipulidae sent by them. Except where indicated to the contrary, the types of the novelties are preserved in my collection.
NEW DESCRIPTIONS OF LARVAE OF FOREST INSECTS: NYCTOBIA, EUFIDONIA (LEPIDOPTERA, GEOMETRIDAE)
Authors: McGuffin W. C.
Published: January 2010

Amongst the several species of greon loopers found on Canadian coniferous trees in July are two which, to the naked eye, appear somewhat similar. These are Nyctobia limitaria Wlk. and Eufidonia notataria Wlk. Larvae of these two, along with that of the other Nearctic species of Eufidonia, i.e. discospilata Wlk., a heath-plant feeder, are herewith described. The larva of E. notataria has been described briefly by Goodell (1, 2).
THE OCCURRENCE IN NORTH AMERICA OF THE EUROPEAN STAPHYLINID DELEASTER DICHROUS GRAV.
Authors: MacNay C. G.
Published: January 2010

Recent tests of the effects of a 5 per cent DDT dust on various household insects in the basement of a cafeteria near Eastview, Ontario, resulted in a high mortality among several species of Coleoptera occurring there.
NEW NORTH AMERICAN GEOMETRIDAE WITH NOTES, IV
Authors: McDunnough J.
Published: January 2010

Allied to pontiaria Tayl. in male genitalia bat the numerous teeth along the chitinized costa of the clasper much smaller; the strong excavation of this section found in pontiaria is much reduced in the present species and the membranous portion of the clasper is subequal in length to that of the chitinzed area and not considerably shorter. The whole genitalia are noticeably smaller.
LARVAE OF SOME CANADIAN GEOMETRIDS
Authors: McGuffin W. C.
Published: January 2010

Larval descriptions are useful, but if these same descriptions can be satisfactorily tied together by an artificial key, their value is greatly enhanced. Herewith a key is presented for the determination of mature geometrid larvae on which notes have been published in this periodical since 1941.
STUDIES IN THE MALACHIIDAE
Authors: Marshall M. Y.
Published: January 2010

The recent, acquisition of material in the present subfamily Malachiinae has brought to light several new species, as well as afforded an opportunity for observations on other species in the group that are believed to be worthy of recording.
A NEW SPECIES OF DELPHACODES FROM ALBERTA (FULGOROIDEA)
Authors: Metcalf Z. P.
Published: January 2010

Among the many interesting Homoptera which have been sent to me by Professor E. H. Strickland is a series of an unusual Delphacodes collected at Cooking Lake, Alberta. This distinct little species is described at this time in order to stimulate further collection of these inconspicuous but interesting insects from western Canada, where there must be many more new species as yet unrecorded.
NEW DESCRIPTIONS OF LARVAE OF FOREST INSECTS: SEMIOTHISA, DYSMIGIA (LEPIDOPTERA, GEOMETRIDAE)
Authors: McGuffin W. C.
Published: January 2010

Some geometrid larvae exhibit dimorphism in colour. Several interesting observations of this phenomenon were made during the study of larvae of the species, Semiothisa sexmaculata Pack., Dysmigia loricaria Evers., and Nepytia canosaria Wlk.

In these observations, certain points stand out. Although larvae of S. sexmaculata are always green until they reach the last instar, both brown and green phases occur in that stage. After the last larval moult, a transition period of approximately two days is required for larvae to acquire the colouring of the brown phase. On the other hand, specimens of D. loricaria may pass their entire larval period in either a brown or a green phase (as do the larvae of N. canosaria, descriptions of which may be found in Can. Ent. 75: 186-189) or they may change (as do some larvae of S. sexmaculata) from the green phase to the brown phase in the last larval instar. For Dysmigia larvae to undergo this transformation, approximately three days are required.
LABORATORY ASSESSMENT OF ORGANIC INSECTICIDES FOR CONTROL OF CERTAIN LEPIDOPTEROUS LARVAE
Authors: Brown A. W. A. , Hopewell W. W. , Wenner B. J. , McDonald H.
Published: January 2010

Recent developments in the insecticide field have made available to the economic entomologist a wide variety of chemical compounds which exhibit high toxicity to certain species or groups of insects. The pyrethrins rotenone and nicotine, reinforced by the dinitro compounds and thiocyanates of prewar times, have now been joined by the chlorinated aromatic hydro-carbons: DDT, gammexane, chlordane and toxapohene.
OBSERVATIONS ON THE EMERGENCE OF PHOROCERA HAMATA A. AND W. (TACHINIDAE) FROM SAWFLY COCOONS
Authors: Baldwin W. F. , Coppel H. C.
Published: January 2010

The literature dealing with the methods by which the larvae and adults of tachinid parasites escape from sawfly cocoons indicates in certain cases that only circumstantial evidence is available concerning the formation of the exit hole in nature. Among these parasites a large number leave their hosts while the sawflies are in the larval stages. The larvae make their escape as mature maggots through a hole gnawed in the soft host integument. A few species, however, continue to develop within the host after the host cocoon has been formed. Some of these cut through the cocoon as mature larvae and later pupate in a suitable medium, while others form their puparial cases within the host cocoons, the adults later escaping from both the puparial cases and from the sawfly cocoons. The only escape mcchanism possessed by an adult tachinid fly consists ot the alternate expansion and contraction of the ptilinum against the surrounding wall. Since the cocoon is usually quite tough and leathery it is essential that some provision be made in advance to enable the adult to escape. This seemed to occur with adults of Phorocera hamata A. and W. which emerge from their puparia within the host cocoon and then escape from the cocoon proper, leaving a characteristic round cap which appears to have been cut (Fig. 1, C). The method of formation of this cap has been the subject of discussion by several authors.
A Study of the Male Hypopygia of the Species of Tubifera (Syrphidae, Diptera) That Occur North of Mexico
Authors: Bean James L.
Published: January 2010

During a study of the species of Tubifera known to occur in Maine the author noted that certain specific key characters were not always constant, and often a long series of several species was required before identification was certain. In search of taxonomic characters to supplement those given in literature the author extracted and compared the male hypopygia of several known species of Tubifera. It soon became apparent that this structure deserved more emphasis in the taxonomy of this interesting genus.
Culture of Entomophagous Insects
Authors: Flanders Stanley E.
Published: January 2010

“When we consider what immense services hymenopteron insects render to agriculture, it is surprising that those states that have spent already such large sums of money to fight noxious insects and that have lost still more through their depredation have never tried to raise ichneumon flies by the million and let them loose wherever there are any insect pests to destroy.” Felix Gillet., Hort. Comm., El Dorado, California, 1882.
A Critique on Insect Chemical Control Methods
Authors: Pickett A. D.
Published: January 2010

While it is said, with some truth, that man's greatest conflict is with man, and although that conflict shows little indication of soon abating, he faces other conflicts which are also very real and immediate. Man is one of the many competing species in the world's interdependent fauna and his very existence is inextricably interwoven with that of the other animals in his environment. Man's progress may be measured largely by the degree to which he has been able to shape his environment to meet his needs and few would deny that in so altering his environment he has made human existence more tolerable and consequently, more satisfying. At least few of us would elect to return to a more primitive stage of human history.
Fluorescence Microscopy: A Concise Guide to Current Imaging Methods
Authors: Combs Christian A.
Published: January 2010

The field of fluorescence microscopy is rapidly growing, providing ever increasing imaging capabilities for cell and neurobiologists. Over the last decade, many new technologies and techniques have been developed which allow for deeper, faster, or higher resolution imaging. For the non‐expert microscopist, it can be difficult to match the best imaging technique to the biological question to be examined. Picking the right technique requires a basic understanding of the underlying imaging physics for each technique, as well as an informed comparison and balancing of competing imaging properties in the context of the sample to be imaged. This unit provides concise descriptions of a range of commercially available imaging techniques and provides a tabular guide to choosing among them. Techniques covered include structured light, confocal, total internal reflection fluorescence (TIRF), two‐photon, and stimulated emission depletion (STED) microscopy. Curr. Protoc. Neurosci. 50:2.1.1‐2.1.14. © 2010 by John Wiley & Sons, Inc.
Loading Neurons with Dextran‐Conjugated Calcium Indicators in Intact Nervous Tissue
Authors: Delaney Kerry R.
Published: January 2010

Dextran‐conjugated Ca2+ indicators are retained well in neurons for many days following loading in intact or semi‐intact brain tissue. Methods for loading neurons, as well as discussion of the unique properties of dextran‐conjugated dyes which need to be considered for their use, are presented. Curr. Protoc. Neurosci. 50:2.5.1‐2.5.11. © 2010 by John Wiley & Sons, Inc.
Gene Transfer into Rat Brain Using Adenoviral Vectors
Authors: Puntel Mariana , Kroeger Kurt M. , Sanderson Nicholas S.R. , Thomas Clare E. , Castro Maria G. , Lowenstein Pedro R.
Published: January 2010

Viral vector–mediated gene delivery is an attractive procedure for introducing genes into the brain, both for purposes of basic neuroscience research and to develop gene therapy for neurological diseases. Replication‐defective adenoviruses possess many features which make them ideal vectors for this purpose—efficiently transducing terminally differentiated cells such as neurons and glial cells, resulting in high levels of transgene expression in vivo. Also, in the absence of anti‐adenovirus immunity, these vectors can sustain very long‐term transgene expression within the brain parenchyma. This unit provides protocols for the stereotactic injection of adenoviral vectors into the brain, followed by protocols to detect transgene expression or infiltrates of immune cells by immunocytochemistry or immunofluorescence. ELISPOT and neutralizing antibody assay methodologies are provided to quantitate the levels of cellular and humoral immune responses against adenoviruses. Quantitation of adenoviral vector genomes within the rat brain using qPCR is also described. Curr. Protoc. Neurosci. 50:4.24.1‐4.24.49. © 2010 by John Wiley & Sons, Inc.
Generation of Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) Using Directed Molecular Evolution
Authors: Pei Ying , Dong Shuyun , Roth Bryan L.
Published: January 2010

G protein–coupled receptors (GPCRs) and their signal transductions are important for both physiological and pathological processes in neuron systems. Neuronal GPCRs activated by synthetic ligands have been created by designed mutagenesis for studying their functions and signal pathways. However, these engineered GPCRs have problems, such as their high constitutive activity. To overcome this drawback, a new generation of receptors termed designer receptors exclusively activated by designer drugs (DREADDs), have been designed. DREADDs are exclusively activated by synthetic ligands, but are insensitive to their endogenous ligand and have no constitutive activity, which provides the ability to selectively modulate signal transduction of certain GPCRs in vitro and in vivo. This protocol provides detailed instructions for creating DREADDs using directed molecular evolution. The procedures to generate DREADDS include GPCR functional expression in yeast, mutant GPCR library generation, and high‐throughput yeast screening. These methods are general and suitable for any GPCRs that can be functionally expressed in yeast. Curr. Protoc. Neurosci. 50:4.33.1‐4.33.25. © 2010 by John Wiley & Sons, Inc.
RNA‐Seq: A Method for Comprehensive Transcriptome Analysis
Authors: Nagalakshmi Ugrappa , Waern Karl , Snyder Michael
Published: January 2010

A recently developed technique called RNA Sequencing (RNA‐Seq) uses massively parallel sequencing to allow transcriptome analyses of genomes at a far higher resolution than is available with Sanger sequencing‐ and microarray‐based methods. In the RNA‐Seq method, complementary DNAs (cDNAs) generated from the RNA of interest are directly sequenced using next‐generation sequencing technologies. The reads obtained from this can then be aligned to a reference genome in order to construct a whole‐genome transcriptome map. RNA‐Seq has been used successfully to precisely quantify transcript levels, confirm or revise previously annotated 5′ and 3′ ends of genes, and map exon/intron boundaries. This unit describes protocols for performing RNA‐Seq using the Illumina sequencing platform. Curr. Protoc. Mol. Biol. 89:4.11.1‐4.11.13. © 2010 by John Wiley & Sons, Inc.
Endoglycosidase and Glycoamidase Release of N‐Linked Glycans
Authors: Freeze Hudson H. , Kranz Christian
Published: January 2010

Nearly all proteins entering the lumen of the endoplasmic reticulum (ER) become glycosylated en route to a cellular organelle, the plasma membrane, or the extracellular space. Many glycans can be attached to proteins, but the most common are the N‐linked glycans (oligosaccharides). These chains are added very soon after a protein enters the ER, but they undergo extensive remodeling (processing), especially in the Golgi. Processing changes the sensitivity of the N‐glycan to enzymes that cleave entire sugar chains or individual monosaccharides, which also changes the migration of the protein on SDS gels. These changes can be used to indicate when a protein has passed a particular subcellular location. This unit details some of the methods used to track a protein as it trafficks from the ER to the Golgi toward its final location. Curr. Protoc. Mol. Biol. 89:17.13A.1‐17.13A.25. © 2010 by John Wiley & Sons, Inc.
Galaxy: A Web‐Based Genome Analysis Tool for Experimentalists
Authors: Blankenberg Daniel , Kuster Gregory Von , Coraor Nathaniel , Ananda Guruprasad , Lazarus Ross , Mangan Mary , Nekrutenko Anton , Taylor James
Published: January 2010

High‐throughput data production has revolutionized molecular biology. However, massive increases in data generation capacity require analysis approaches that are more sophisticated, and often very computationally intensive. Thus, making sense of high‐throughput data requires informatics support. Galaxy (http://galaxyproject.org) is a software system that provides this support through a framework that gives experimentalists simple interfaces to powerful tools, while automatically managing the computational details. Galaxy is distributed both as a publicly available Web service, which provides tools for the analysis of genomic, comparative genomic, and functional genomic data, or a downloadable package that can be deployed in individual laboratories. Either way, it allows experimentalists without informatics or programming expertise to perform complex large‐scale analysis with just a Web browser. Curr. Protoc. Mol. Biol. 89:19.10.1‐19.10.21. © 2010 by John Wiley & Sons, Inc.
Chromatin Interaction Analysis Using Paired‐End Tag Sequencing
Authors: Fullwood Melissa J. , Han Yuyuan , Wei Chia‐Lin , Ruan Xiaoan , Ruan Yijun
Published: January 2010

Chromatin Interaction Analysis using Paired‐End Tag sequencing (ChIA‐PET) is a technique developed for large‐scale, de novo analysis of higher‐order chromatin structures. Cells are treated with formaldehyde to cross‐link chromatin interactions, DNA segments bound by protein factors are enriched by chromatin immunoprecipitation, and interacting DNA fragments are then captured by proximity ligation. The Paired‐End Tag (PET) strategy is applied to the construction of ChIA‐PET libraries, which are sequenced by high‐throughput next‐generation sequencing technologies. Finally, raw PET sequences are subjected to bioinformatics analysis, resulting in a genome‐wide map of binding sites and chromatin interactions mediated by the protein factor under study. This unit describes ChIA‐PET for genome‐wide analysis of chromatin interactions in mammalian cells, with the application of Roche/454 and Illumina sequencing technologies. Curr. Protoc. Mol. Biol. 89:21.15.1‐21.15.25. © 2010 by John Wiley & Sons, Inc.
Mapping Networks of Protein‐Mediated Physical Interactions Between Chromatin Elements
Authors: Tiwari Vijay K. , Baylin Stephen B.
Published: January 2010

Understanding of transcriptional regulation has advanced in recent years in part due to development of technologies which allow determination of physical proximities between interacting chromatin regions at a resolution beyond that offered by conventional microscopy techniques. However, these methods do not specifically identify the protein component(s) that might mediate such interactions. This unit provides a detailed protocol for Combined 3C‐ChIP‐Cloning (6C) technology, which combines multiple techniques to simultaneously identify physical proximities between chromatin elements as well as the proteins that mediate these interactions. The unit further explores how the 6C assay can be combined with other techniques for a complete, cell‐type‐specific mapping of all inter‐ and intrachromosomal interactions mediated by specific proteins. Thus, the 6C assay provides a useful tool to address the role of specific proteins in nuclear organization and to advance our understanding about the relation of chromatin higher‐order organization and transcriptional regulation. Curr. Protoc. Mol. Biol. 89:21.16.1‐21.16.13. © 2010 by John Wiley & Sons, Inc.
Methylation‐Sensitive Single‐Molecule Analysis of Chromatin Structure
Authors: Miranda Tina B. , Kelly Theresa K. , Bouazoune Karim , Jones Peter A.
Published: January 2010

Methylation‐sensitive single‐molecule analysis of chromatin structure is a high‐resolution method for studying nucleosome positioning. As described in this unit, this method allows for the analysis of the chromatin structure of unmethylated CpG islands or in vitro–remodeled nucleosomes by treatment with the CpG‐specific DNA methyltransferase SssI (M.SssI), followed by bisulfite sequencing of individual progeny DNA molecules. Unlike nuclease‐based approaches, this method allows each molecule to be viewed as an individual entity instead of an average population. Curr. Protoc. Mol. Biol. 89:21.17.1‐21.17.16. © 2010 by John Wiley & Sons, Inc.
Overview of Very Small Embryonic‐Like Stem Cells (VSELs) and Methodology of Their Identification and Isolation by Flow Cytometric Methods
Authors: Zuba‐Surma Ewa K. , Ratajczak Mariusz Z.
Published: January 2010

The protocols presented here describe the procedures employed to identify and isolate very small embryonic‐like stem cells (VSELs) using flow cytometric technologies including fluorescence‐activated cell sorting (FACS). We describe the recommended steps in detail for their successful identification and isolation from adult tissues. These protocols were initially established to isolate such cells from murine bone marrow (BM) and human cord blood (CB) and may also be employed to isolate these primitive cells from other adult organs and embryonic tissues. Here, we focus on some critical parameters/key points required for the successful identification and purification of these rare cells by employing classical flow cytometry. In the last part of this unit, we also discuss a novel flow cytometric tool, ImageStream, an imaging flow cytometer, which allows better identification and morphological analysis of sorted cells. Curr. Protoc. Cytom. 51:9.29.1‐9.29.15. © 2010 by John Wiley & Sons, Inc.
Use of Flow Cytometric Methods to Quantify Protein‐Protein Interactions
Authors: Blazer Levi L. , Roman David L. , Muxlow Molly R. , Neubig Richard R.
Published: January 2010

A method is described for the quantitative analysis of protein‐protein interactions using the flow cytometry protein interaction assay (FCPIA). This method is based upon immobilizing protein on a polystyrene bead, incubating these beads with a fluorescently labeled binding partner, and assessing the sample for bead‐associated fluorescence in a flow cytometer. This method can be used to calculate protein‐protein interaction affinities or to perform competition experiments with unlabeled binding partners or small molecules. Examples described in this protocol highlight the use of this assay in the quantification of the affinity of binding partners of the regulator of G‐protein signaling protein, RGS19, in either a saturation or a competition format. An adaptation of this method that is compatible for high‐throughput screening is also provided. Curr. Protoc. Cytom. 51:13.11.1‐13.11.15. © 2010 by John Wiley & Sons, Inc.
Metamaterials
Authors: Iyer Ashwin K. , Eleftheriades George V.
Published: January 2010

The sections in this article are

Introduction

Background

Transmission‐Line Theory ofLHMedia

Metamaterial Implementations

Metamaterial Applications

Beyond Left‐Handedness
Knowledge‐Based Systems
Authors: Hudson Donna L.
Published: January 2010

The sections in this article are

Introduction

History

Knowledge‐Based Systems In Other Domains

Knowledge Acquisition and Validation

New Methodologies
Real‐Time Embedded Software
Authors: Edwards Stephen A.
Published: January 2010

Real‐time embedded software is responsible for observing and controlling systems in the physical world. Because of this role, it has timeliness requirements above and beyond that of traditional software: when it produces results is often as important the results themselves. The real‐time challenge, compounded with the usual difficulties of producing correct, complex software systems, makes producing real‐time embedded software all the more difficult.

Such software runs on digital computing hardware, is usually structured following principles from real‐time scheduling, and often implements algorithms derived from control theory. Effective development of such software requires knowledge of these three fields.

Low‐level concerns such as memory‐mapped I/O, polling, and interrupts are discussed first, followed by a discussion of how to simulate concurrency and PThreads. An introduction to real‐time scheduling theory follows, including a discussion of the rate‐monotonic approach. The article ends with a discussion of real‐time operating systems and the future of real‐time embedded software.
Bode Plots
Authors: Schubert Thomas F.
Published: January 2010

The sections in this article are

Introduction

Bode Plots of the Factors
Hair Cells
Authors: Marcotti Walter , Masetto Sergio
Published: January 2010

Hair cells are the sensory receptors in the inner ear that detect sound and head motion to begin the processes of hearing and balance control. The defining feature of hair cells is the hair bundle, the transduction organelle protruding from their apical surface composed of ordered arrays of stereocilia. Mechanical deflection of the hair bundle, normally induced by physiological stimuli, increases the open probability of mechanically gated cation channels located at the tip of stereocilia. The resulting depolarizing inward current generates a receptor potential. The information encoded in this electrical response is transmitted to the auditory or vestibular afferent nerve fibres via the Ca
2+
‐induced release of neurotransmitter from the hair cell's basal pole. In this way sensory information is relayed to the brain enabling us to perceive sound and maintain balance. In mammals, hair cell loss causes irreversible balance and hearing impairment because these sensory cells show very little or no regenerative ability.

Key concepts:

Hair cells are the sensory receptors of both the auditory and vestibular systems in vertebrates.

Inner ear hair cells derive their name from the stereocilia that protrude from their apical surface, which are deflected as a result of sound (cochlea) or head movement (vestibular organs) and initiate the sensory transduction process.

Cochlear hair cells convert acoustic stimuli into electrical activity that allows the brain to perceive the sensation of hearing.

Vestibular hair cells inform the central nervous system of head position and movements.

The hair bundle, the defining feature of all hair cells, is the mechano‐receptive organelle directly responsible for mechano‐transduction, i.e. for converting the natural stimulus into a neural signal.

In mammals, hair cell loss due to acoustic over‐stimulation, ototoxic drugs, ageing and genetic defects is irreversible, leading to a permanent loss of function.
Substance P
Authors: Bremer Andrew A , Leeman Susan E
Published: January 2010

Substance P (SP) is an 11 amino acid bioactive peptide that causes diverse biological effects in the central nervous system (CNS), as well as in the cardiovascular, respiratory, gastrointestinal, immune and autonomic nervous systems. The principal actions of SP are mediated through its functional interaction with the neurokinin‐1 (NK‐1) receptor, which exists in two isoforms. Investigators are currently focusing on the structure–activity relationships between SP and the different NK‐1 receptor isoforms, as well as the signal transduction pathways involved in the many SP‐mediated responses. In addition, nonpeptide antagonists of SP have been and are continuing to be developed for clinical use. This article summarizes the basic biology of SP and its interaction with the NK‐1 receptor, highlights the role of SP and the neurokinin receptors in clinical disease states and then concludes with describing new exciting areas of SP‐related research.

Key Concepts:

Substance P is an 11 amino acid bioactive peptide that induces diverse biological responses.

The principal receptor for substance P is the neurokinin‐1 receptor, which exists in two isoforms.

Antagonists of substance P and the neurokinin‐1 receptor have been and are continuing to be developed for clinical use.

Research in the area of substance P and the neurokinin‐1 receptor continues to provide more insight into the importance of these molecules in states of human health and disease.
Eukaryotic Replication Protein A
Authors: Humphreys Troy D , Wold Marc S
Published: January 2010

Replication protein A
(
RPA
) is a single‐stranded
deoxyribonucleic acid
(DNA)‐binding protein required for cellular DNA metabolism.
RPA
is a heterotrimeric complex composed of 70, 32 and 14 kDa subunits, commonly referred to as
RPA1
,
RPA2
and
RPA3
. Homologous complexes are found in all eukaryotes.
RPA
binds single‐stranded DNA with high‐affinity and low‐sequence specificity. In addition,
RPA
interacts specifically with, and modulates the activity of, many proteins required for DNA metabolism in cells.
RPA
primarily functions in the cell to stabilize single‐stranded DNA.
RPA
activity is essential for DNA replication, DNA repair and recombination. In addition,
RPA
is involved in the cellular DNA damage response. Disruption of
RPA
activity or of the activity of
RPA
‐interacting proteins is associated with many human diseases, including cancer.

Key Concepts:

Replication protein A (RPA) is the major eukaryotic single‐stranded DNA‐binding protein.

RPA is a heterotrimeric protein complex with a flexible structure; each subunit is composed of one or more DNA‐binding domains that are linked or flanked by flexible linkers and regulatory regions.

RPA participates in many cellular processes essential for life, including DNA replication and repair by binding to single‐stranded DNA intermediates and interacting with proteins required for these processes.
Stamens
Authors: Smyth David R
Published: January 2010

Stamens are the male reproductive organs of flowers. Typically, a stamen is made up of a pollen‐producing anther borne on a filament. Pollen arises in four sporogenous regions within the anther. The sporogenous cells are surrounded by nutritive support cells, the tapetum. Mature pollen is released from the anther by dehiscence of the anther wall. The developmental identity of the stamen is conferred by the combined action of MADS transcription factors. Subsequent cellular differentiation is under the control of a cascade of other transcription factor and signalling genes now being identified. Male sterility is useful in plant breeding, and may arise by loss‐of‐function of these genes, as well as by defects in mitochondria (cytoplasmic male sterility). Differences in the structure of stamens are associated with the types of pollinators involved. Stamens evolved from the microsporangia‐bearing organs of early seed plants.

Key Concepts:

Pollen is generated in stamens, the male reproductive organs of flowers.

Pollen arises from sporogenous cells within the anthers.

Sporogenous cells are surrounded by the tapetum, a nutritive layer of support cells.

Stamen identity is conferred by the action of MADS transcription factors controlling B, C and SEPALLATA function.

The transcription factor SPOROCYTELESS is required for generation of the sporogenous cells.

Other transcription factors and signalling kinases are required for the entire developmental cascade and are now being identified.

Mature pollen is released by dehiscence of the anther wall through slits called stomia.

The structure of stamens in a species is closely adapted to the specific pollinators involved for that species.

Pollen is also generated in nonflowering seed plants (gymnosperms and various extinct lineages), but it arises in leaf‐like organs (microsporophylls) rather than stamens.
Complement: Deficiency Diseases
Authors: Blom Anna M
Published: January 2010

The complement system is crucial for defence against pathogens, removal of unwanted materials such as dying cells or immune complexes as well as for development of adaptive immune responses. Genetically determined deficiencies of components of the complement system are usually relatively rare but they result in many severe diseases such as an increased susceptibility to recurrent, severe infections, autoimmune disorders (systemic lupus erythematosus), glomerulonephritis including membranoproliferative glomerulonephritis type II, paroxysmal nocturnal haemoglobinuria or angioedema. Recently, mutations and polymorphisms in complement proteins, particularly complement inhibitors, have been associated with atypical haemolytic uremic syndrome and age‐related macular degeneration. The elucidation of the pathophysiological basis for the different clinical presentations of complement‐deficient individuals has contributed to a better understanding of the physiological role of complement in normal individuals as well as to the development of emerging therapies.

Key Concepts

The majority of complement deficiencies are rare but cause severe diseases.

Deficiencies of the early components of the classical pathway (C1q/r/s, C4, C2) predispose to systemic lupus erythematosus.

The majority of complement deficiencies predispose to infections either withNeisseriaspecies (deficiency of Factor D, Properdin, C5, C6, C7, C8, C9) or Gram‐positive bacteria (C1q/r/s, mannose‐binding lectin, C2, C4, C3, factor I).

Mutations in C1q/r/s, C2, C4, C3 and factor I can cause glomerulonephritis.

Paroxysmal nocturnal haemoglobinuria is caused by deficiency of complement inhibitors CD55 and CD59.

Hereditary angioedema is found in patients with dysfunctional C1‐inhibitor.

Atypical haemolytic uremic syndrome is mainly associated with mutations and polymorphisms in complement inhibitors.

Age‐related macular degeneration is associated with polymorphisms in complement inhibitor factor H.
Cerebrospinal Fluid and its Abnormalities
Authors: Chan Michael , Amin‐Hanjani Sepideh
Published: January 2010

Cerebrospinal fluid bathes the central nervous system, providing mechanical support and maintaining a stable environment. Alterations in its production, circulation or absorption result in a pathological condition termed hydrocephalus, characterized by an accumulation of fluid within the cranium, usually in the ventricles of the brain. Similarly, aberrations in the circulation or absorption of cerebrospinal fluid around the spinal cord can create a condition termed syringomyelia. Hydrocephalus can be life‐threatening due to increased intracranial pressure. The most common causes of hydrocephalus are related to obstruction of cerebrospinal fluid flow. In the pediatric population, causes of hydrocephalus include developmental abnormalities, infectious aetiologies, mass lesions, and haemorrhage. Causes of high pressure hydrocephalus in adulthood are primarily related to haemorrhage and tumour, although idiopathic normal pressure hydrocephalus (NPH) is also encountered. The management of hydrocephalus and syringomyelia relies primarily on neurosurgical interventions such as insertion of shunt devices.

Key Concepts

CSF provides a mechanical support for the brain and spinal cord.

Overproduction of CSF, obstruction to CSF flow or impaired absorption of CSF may lead to disease states such as hydrocephalus or syringomyelia.

CSF diversion by means of shunting with catheter implantation or fenestration of anatomic barriers is the primary treatment for diseases of CSF disturbances.