bluuebunny/crossref_metadata_2025 huggingface dataset:
Vargas, Eisenhower e a Questão Guatemalteca, 1954: Uma Análise de Política Externa
Authors: Avila Carlos Federico Domínguez
Published: May 2022

RESUMO O artigo explora a formulação e implementação da política brasileira diante da denominada questão guatemalteca, em 1954. O artigo é resultado de pesquisa histórico-política no Arquivo do Ministério das Relações Exteriores. Foram considerados aproximadamente 500 documentos desclassificados (secretos e confidenciais). No contexto da história da Guerra Fria, o artigo discute a convergência entre a política externa brasileira e a estadunidense em relação à questão guatemalteca, principalmente em três foros: uma projetada Reunião de Consulta dos Ministros das Relações Exteriores do continente americano; no Conselho de Segurança da Organização das Nações Unidas; e na Comissão Interamericana de Paz, vinculada à Organização dos Estados Americanos. As fontes primárias consultadas sugerem que existiu uma predisposição do lado brasileiro em acompanhar as iniciativas diplomáticas do governo dos Estados Unidos em relação à questão guatemalteca. Contudo, não é possível documentar se havia conhecimento, do lado brasileiro, acerca da operação encoberta PBSUCCESS. Daí que poderia ter acontecido alguma manipulação dos diplomatas e políticos brasileiros.
The Flight of the Junky: Existential Posthumanism and Immanent Life in Early Burroughs
Authors: Hoffman Todd
Published: May 2022

William Burroughs early book Junky is generally separated from his later experimental fiction.  Stylistically it accords much more to realism than the postmodern aleatory method he later innovated.  However, Burroughs’ preoccupation with resisting all forms of subjectification, his disenchantment with bourgeois life and his simultaneous literal and tropic use of addiction as a form of flight from powers of normalization and conformity are strongly present in this early work.  This paper explores Junky on three fronts.  First, it shows the novel as an elaboration of a posthumanist existentialism by emphasizing the materiality of the body through Burroughs’ explanation of the physiological mechanisms of addiction.  Through this existentialist posthumanism, the novel critically responds to Sartrian existentialism, which was so fashionable at the time of Burroughs’ writing, and repudiates the Jeffersonian idealization of the transcendental subject and its middle class figurations.  The emphasis on the material body simultaneously challenges post-structuralist renderings of Burroughsian readings.  This leads to a conception of strategies of flight from all forms of conformity by utilizing Deleuze and Guattari’s notion of the Body without Organs and immanent Life.  Junk is a vehicle of flight and self-affirmation, a means of highly individualized, libertarian modes of subjective deterritorialization.  Addiction and habitual use are not mere uncontrolled thirsts, but forms of actualizing a wholly detached social and independent individual.  But the danger of junk lies in its reterritorializing of the body through new assemblages of need and dependence, leading the protagonist to ultimately seek a different mode of escape.  Junk illuminates our posthuman existential condition and leads Burroughs to seek new experimental forms of aesthetic expression.
POWER CONSUMPTION FORECASTING BASED ON FULLY CONNECTED FEED-FORWARD NEURAL NETWORKS
Authors:   National Research Technological University «MISIS», Lyakhomsky Alexander , Shadrin Andrei ,   National Research Technological University «MISIS»
Published: May 2022

The relevance The relevance of electricity consumption forecasting on the basis of fully connected feed-forward neural networks (FNN) to improve the validity of applications for electricity is considered. Aim of research Synthesis of predictive model of electricity consumption in the form of four-layer fully connected feed-forward neural network, linking the volume of production and the predicted electricity consumption is performed. Research methods The algorithm of the predictive model development includes: formation and initial statistical processing of initial data; determination of FNN structure hyperparameters — total number of layers, number of neurons in layers, activation function, training rate coefficient; selection of optimization method; training, checking model adequacy. Results The analytical expression for the description of the forecast model based on FNN is given. The synthesized forecast model makes it possible to increase the validity of electric power applications of enterprises.
MICROPROCESSOR SYSTEM FOR AUTOMATIC CONTROL OF AIRCRAFT FIRE PROTECTION MEANS
Authors:   Ufa State Aviation Technical University, Kumertau, Zaitseva Alina , Dudayev Nikolai ,   Ufa State Aviation Technical University, Kumertau, Shcherbakov Konstantin ,   Ufa State Aviation Technical University, Kumertau
Published: May 2022

Relevance The ever-increasing requirements for the safety of the use of aviation technology are inextricably linked with the problem of providing fire protection for aircraft, both military and civilian. The complexity of the problem of ensuring the fire safety of flights is associated with an increase in the intensity of the use of aviation equipment and the expansion of the range of functional tasks performed by it. The resulting complication of on-board equipment and an increase in the number of energy-intensive devices creates the prerequisites for the occurrence of fires on board the aircraft. At the same time, the remoteness of places where fires are possible, the variety of causes leading to fires, as well as the ambiguity of the conditions for the onset and spread, increase the likelihood of equipment failures, but also complicate the crew's activities. The purpose of the study is to implement a prospective aircraft fire protection system that will increase the effectiveness of existing fire extinguishing equipment. The relevance of this research project lies in the creation of an aircraft fire-fighting system that will provide timely detection of overheating/fire in the nacelles of the main power unit, in the compartments of the auxiliary power unit, baggage and cargo compartments and aircraft toilets; reliability of information from fire detection and elimination systems; increase the effectiveness of existing firefighting equipment. Aim of research The aim of the study is to develop a future aircraft fire protection system that will increase the effectiveness of existing fire extinguishing equipment. The objectives of the research project are: 1. Choosing a hardware complex for electronic indication and signaling of the aircraft fire-fighting system; 2. Integration of the fire protection complex into the general aircraft equipment control system. Research methods Analysis of modern high-performance aircraft fire protection systems and creation of a promising fire protection system based on the data obtained Results In the course of this research project, a hardware complex for electronic indication and signaling of the aircraft fire system was selected, and the integration of the fire protection complex into the control system of general aircraft equipment was carried out.
RELIABILITY PROBLEMS OF FREQUENCY-CONTROLLED ELECTRIC MOTORS
Authors:   National Research Tomsk Polytechnic University, Kolesnikov Stanislav , Leonov Andrey ,   National Research Tomsk Polytechnic University
Published: May 2022

Relevance Widespread use of semiconductor frequency converters is due to the significant advantages of this technology: smoothness and accuracy of controlling the speed and torque of the electric motor, smooth start and stop; increased energy efficiency; the possibility of remote monitoring, etc. The accumulated operational experience has shown an increased failure rate of stator winding insulation of asynchronous motors operating with pulsed power. The presence of high-frequency overvoltages in the inverter signal can lead to the appearance of partial discharges even in the low-voltage motors windings. It’s leading to accelerated degradation of electrical insulation and premature breakdown. Over the past three decades, a significant amount of information has been accumulated in the technical literature: high-frequency transient models have been created; partialdischarge and corona-resistant test methods have been developed; the most important voltage parameters determining the intensity of insulation aging have been determined; regulatory documentation for testing and selection of insulation systems have been developed. Analysis and systematization of existing results are necessary to determine the directions of further research in the subject. Аim of research Identification of new solutions and trends of insulation reliability windings of lowvoltage electric machines when powered by modulated voltage. Research methods Bibliographic analysis, methods of statistical data processing. Results Analysis of the most relevant works on the reliability of low-voltage winding insulation systems in frequency electric drive systems is carried out, a bibliographic list of these works is given. The main results concerning the overvoltages simulation, insulation testing methods, studying the aging mechanisms of winding polymer insulation with pulsed power supply, determining the characteristics of emerging discharges are shown. The ways of further development aimed at modeling overvoltages, taking into account high-frequency effects, increasing the volume of experiments to estimate the insulation lifetime, studying the conditions for the occurrence of partial discharges, creating a common approach to insulation testing are determined.
Taxon group: Terrestrial Arthropods smaller than 5mm (TSS3) v1
Authors: Pereira da Conceicoa Lyndall , Sivell Olga , Sivess Laura , Fletcher Chris , R. Broad Gavin , Crowley Liam , Januszczak Inez
Published: May 2022

This is part of the collection "DToL Taxon-specific Standard Operating Procedure (SOP) for the Terrestrial and Freshwater Arthropods Working Group". The SOP collection contains guidance on how to process the various terrestrial and freshwater arthropod taxa within the scope of the Darwin Tree of Life project. The guidance specifically refers to the tissue samples needed for DNA barcoding (which takes place at the Natural History Museum (NHM)) and outlines the dissected tissues required for whole genome sequencing (WGS), which takes place at the Wellcome Sanger Institute . Every specimen is submitted for DNA barcoding first before potentially being sent to the Wellcome Sanger institute. TSS3 = Taxon specific SOP for Terrestrial Arthropoda smaller than 5mm Note: For genome sequencing, anything smaller than a lentil (5mm), or estimated to be around 5mm, should be frozen whole (live) after photography. There are numerous species that fit under this category. Tissue or extract from Sanger will be used for DNA barcoding. Including: Arachnidae (especially Acari, Pseudoscorpiones), Amphipoda (under 10mm), Collembola, Diplura, Protura, Symphyla, Pauropoda and terrestrial insects (incl. terrestrial and aquatic life stages of freshwater taxa) smaller than 5mm: Archaeognatha, Phthiraptera, Psocoptera, Siphonaptera, Strepsiptera, Thysanoptera, Zygentoma, also small species from remaining orders, e.g. Blattodea (‘Isoptera’), Coleoptera, Diptera, Hymenoptera, Lepidoptera, Mecoptera (Boreidae), Hemiptera.
DToL Taxon-specific Standard Operating Procedure for the Terrestrial and Freshwater Arthropods Working Group v1
Authors: Pereira Lyndall , Sivell Olga , Sivess Laura , Fletcher Chris , R. Broad Gavin , Crowley Liam , Januszczak Inez
Published: May 2022

This Standard Operating Procedure (SOP) collection contains guidance on how to process the various terrestrial and freshwater Arthropod taxa within the scope of the Darwin Tree of Life (DToL) project. The guidance specifically refers to the tissue samples needed for DNA barcoding (which takes place at the Natural History Museum (NHM)) and outlines the dissected tissues required for whole genome sequencing (WGS), which takes place at the Wellcome Sanger Institute . Every specimen is submitted for DNA barcoding first before potentially being sent to the Wellcome Sanger Institute.
[RETRACTED] Chemically Labile Linkers v1
Authors: Sciences BOC
Published: May 2022

[RETRACTED]

The chemically labile linkers, which include acid-cleavable linkers and disulfide linkers, are extensively applied to ADCs due to their ability to undergo fracture, increasing the acidity of the endosomal-lysosomal pathway, as well as the concentration of glutathione inside cells. BOC Scienceshas developed highly integrated approaches that can conjugate antibodies and payload via a linker to compose ADC-related products quickly. We are capable of completing small (mg) and large (g) quantities of ADC small molecule products and transferring these technologies to a bulk manufacturing plant. To ensure quality, all final products will be thoroughly analyzed by our advanced technologies, including mass spectrometry and chromatography.
NEBNext® Immune Sequencing Kit (Human)(E6320) v1
Authors: not provided maria.matos , England Biolabs New
Published: May 2022

This protocol details NEBNext® Immune Sequencing Kit (Human). https://www.neb.com/-/media/nebus/files/manuals/manuale6320.pdf?rev=1dca310ed12a4e89b322063d31db030d&hash=D4D7E9FBED23CC7448447137DBFF8C0B
Extraction and selection of high-molecular-weight DNA for long-read sequencing from Chlamydomonas reinhardtii v1
Authors: Chaux-Jukic Frédéric , Agier Nicolas , Eberhard Stephan , Xu Zhou
Published: May 2022

Recent advances in long-read sequencing technologies have enabled the complete assembly of eukaryotic genomes from telomere to telomere by allowing repeated regions to be fully sequenced and assembled, thus filling the gaps left by previous short-read sequencing methods. Furthermore, long-read sequencing can also help characterizing structural variants, with applications in the fields of genome evolution or cancer genomics. For many organisms, the main bottleneck is to develop robust methods to obtain high-molecular-weight (HMW) DNA for whole genome sequencing purposes. We developed an optimized protocol to extract DNA suitable for long-read sequencing from the unicellular green alga Chlamydomonas reinhardtii, based on CTAB/phenol extraction followed by a size selection step for long DNA molecules. We provide validation results for the extraction protocol, as well as statistics obtained with Oxford Nanopore Technologies sequencing.
MicroStone: Exploring the capabilities of the Artec Micro in scanning stone tools v1
Authors: Falcucci Armando
Published: May 2022

In this protocol, I describe how to scan small-sized lithics with the high-precision desktop 3D scanner Artec Micro. This scanner is particularly efficient when it comes to scan small objects with a 3D resolution of up to 0.029 mm and a 3D accuracy of 0.01 mm. However, a few slight modifications to the common workflow are needed to obtain accurate 3D meshes of small stone tools such as bladelets, microblades, microflakes. Overall, this scanner represents an efficient and reliable alternative to scan artifacts that cannot be successfully captured by other laser or structured light scanners, such as the Artec Space Spider. If several hundreds of artifacts need to be scanned, a less time-consuming alternative is represented by Micro-CT scanners, using a protocol specifically adapted to this task (i.e., the StyroStone Protocol).
P3v4 (v3 from the original) v4
Authors: not provided maria
Published: May 2022

Lorem ipsum dolor sit amet, consectetuer adipiscing elit. Pellentesque arcu. Phasellus faucibus molestie nisl. Quis autem vel eum iure reprehenderit qui in ea voluptate velit esse quam nihil molestiae consequatur, vel illum qui dolorem eum fugiat quo voluptas nulla pariatur? Suspendisse sagittis ultrices augue. Vestibulum erat nulla, ullamcorper nec, rutrum non, nonummy ac, erat. Nullam justo enim, consectetuer nec, ullamcorper ac, vestibulum in, elit. In convallis. Mauris elementum mauris vitae tortor. Sed ac dolor sit amet purus malesuada congue. Etiam ligula pede, sagittis quis, interdum ultricies, scelerisque eu. Nunc auctor. Nam sed tellus id magna elementum tincidunt. Sed convallis magna eu sem. Quis autem vel eum iure reprehenderit qui in ea voluptate velit esse quam nihil molestiae consequatur, vel illum qui dolorem eum fugiat quo voluptas nulla pariatur? In dapibus augue non sapien.
P4v2 v2
Authors: not provided maria
Published: May 2022

Lorem ipsum dolor sit amet, consectetuer adipiscing elit. Pellentesque arcu. Phasellus faucibus molestie nisl. Quis autem vel eum iure reprehenderit qui in ea voluptate velit esse quam nihil molestiae consequatur, vel illum qui dolorem eum fugiat quo voluptas nulla pariatur? Suspendisse sagittis ultrices augue. Vestibulum erat nulla, ullamcorper nec, rutrum non, nonummy ac, erat. Nullam justo enim, consectetuer nec, ullamcorper ac, vestibulum in, elit. In convallis. Mauris elementum mauris vitae tortor. Sed ac dolor sit amet purus malesuada congue. Etiam ligula pede, sagittis quis, interdum ultricies, scelerisque eu. Nunc auctor. Nam sed tellus id magna elementum tincidunt. Sed convallis magna eu sem. Quis autem vel eum iure reprehenderit qui in ea voluptate velit esse quam nihil molestiae consequatur, vel illum qui dolorem eum fugiat quo voluptas nulla pariatur? In dapibus augue non sapien.
Building plan for a temperature-controlled multi-point stirring incubator v1
Authors: Rades Marvin , Schubert Patrick , Ziegler Maren , Kröckel Martin , Reichert Jessica
Published: May 2022

Multiple point stirrers are expensive lab equipment (usually €1,000–4,000) and are only offered by a handful of manufacturers. More importantly, the maximum volume per stirring point is minimal in multi-point stirrers and allows the use of small containers only (usually 0.25–0.6 L). However, for many applications, larger volumes per stirring point are needed. Here, we present a building plan to construct a multiple-point stirrer (with twelve stirring points) for larger container volumes (0.8–3 L) for a fraction of the cost of a commercial device. The overall material cost for a 12-point stirrer is around ca. €280. The stirrer is built with axial flow fans equipped with cuboid magnets and controlled via a programmable power supply. In addition, a water bath (ca. €50) can be placed above the stirring points to allow controlled temperature regulation and to complement the function of an incubator.
P4nv v4
Authors: not provided maria
Published: May 2022

Lorem ipsum dolor sit amet, consectetuer adipiscing elit. Pellentesque arcu. Phasellus faucibus molestie nisl. Quis autem vel eum iure reprehenderit qui in ea voluptate velit esse quam nihil molestiae consequatur, vel illum qui dolorem eum fugiat quo voluptas nulla pariatur? Suspendisse sagittis ultrices augue. Vestibulum erat nulla, ullamcorper nec, rutrum non, nonummy ac, erat. Nullam justo enim, consectetuer nec, ullamcorper ac, vestibulum in, elit. In convallis. Mauris elementum mauris vitae tortor. Sed ac dolor sit amet purus malesuada congue. Etiam ligula pede, sagittis quis, interdum ultricies, scelerisque eu. Nunc auctor. Nam sed tellus id magna elementum tincidunt. Sed convallis magna eu sem. Quis autem vel eum iure reprehenderit qui in ea voluptate velit esse quam nihil molestiae consequatur, vel illum qui dolorem eum fugiat quo voluptas nulla pariatur? In dapibus augue non sapien.
Imaging of Calcium Dynamics in Vasoactive Intestinal Peptide-expressing Neurons of Enteric Nervous System v1
Authors: Margiotta Joseph , Howard Marthe
Published: May 2022

The protocol was published on behalf of the investigators by the SPARC project team. The University of Toledo Health Science Campus animal care and use committee approved animal care, breeding procedures, and experimental protocols. Animals were housed in an Association of Laboratory animal Care-approved facility with a 12-hour light cycle with food (standard chow) and water ad libitum. One male VIP-GCAmP reporter mouse, age 4 months, was used in this study. The mouse was purchased from the Jackson Laboratories, and all genotyping was performed using primers recommended by the Jackson Laboratories according to their protocols.
Extraction of DNA from Frozen Islet Repository Samples v1
Authors: not provided IIDP-HIPP
Published: May 2022

This Standard Operating Procedure (SOP) provides Human Islet Phenotyping Program (HIPP) procedures for DNA extraction from frozen islet pellets using the Promega Wizard® Genomic DNA Purfication Kit. This SOP defines the assay method used by the Human Islet Phenotyping Program (HIPP) for quantitative and qualitative determination of the Purified Human Pancreatic Islet product, post-shipment, manufactured for use in the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)-sponsored research in the Integrated Islet Distribution Program (IIDP).
Lipid transfer assay v1
Authors: not provided patreece.suen
Published: May 2022

This protocol details how to perform a lipid transfer assay with 3xFLAG-SHIP164Δ901-1099
Rab8a expression and purification v1
Authors: M. Snead David
Published: May 2022

Recombinant Rab8a expression and purification protocol as used by the Leschziner and Reck-Peterson Labs. Original protocol by David Snead. Adapted for protocols.io by Mariusz Matyszewski. Current version as used in Snead, Matyszewski, Dickey et al. 2022.
Purification of Human K560-GFP molecular motor v1
Authors: M. Dickey Andrea
Published: May 2022

K560-GFP purification protocol from the Reck-Peterson Lab based on protocol from Nicholas et al. 2014. Edited for protocols.io by Andrea Dickey and Mariusz Matyszewski.
2D TEM CLEM (Correlative Light Microscopy and Electron Microscopy) v1
Authors: De Camilli Pietro , Wu Yumei
Published: May 2022

This protocol details the general procedure of Correlative Light Microscopy and Electron Microscopy (CLEM) with conventional chemical fixation and 2D Transmission Electron Microscopy (TEM) imaging.
Influenza virus plaque assay v1
Authors: F Baker Steven
Published: May 2022

Determine virus concentration from cell culture, lung homogenates, and more. This protocol measures virus in plaque forming units per ml solution. This protocol differs from classic flu plaque assays by using a semi-solid overlay (Avicel) as opposed to agarose. This is beneficial in avoiding reproducibility issues that can arise from burning monolayers with hot overlay, and is faster to make and aspirate. The negative is that you cannot plaque purify viral isolates as you can with agarose. Theoretically one infectious virion can spawn one plaque by initiating in a cell and infecting neighboring cells. This is the gold standard for virology, and for influenza, the plaque:particle ration is ~ 1:100. Thus there are lots of non/semi-infectious or defective interfering particles that are produced that are not on their own infectious.
Data analysis for PLOSONE manuscript v1
Authors: not provided mguillen , M. Guillen, , Robles, I. Bardes , Cabrera, E. Bordera , Roldán, X. Acebes , C. Bolancé, , D. Jorba, , not provided Moriña,
Published: May 2022

These are the data used in manuscript "Acute Respiratory Infection Rates in Primary Care Anticipate ICU Bed Occupancy During COVID-19 Waves"
Synthesis of colloidal dextran-conjugated superparamagnetic iron nanoparticles (SPIONs) v1
Authors: Hancock-Cerutti William , Kumar Tharkeshwar Arun , M Ferguson Shawn , De Camilli Pietro
Published: May 2022

This method describes the synthesis and usage of dextran-conjugated superparamagnetic iron nanoparticles (SPIONs)
Purification of lysosomes using dextran-conjugated SPIONs v1
Authors: Hancock-Cerutti William , Kumar Tharkeshwar Arun , M Ferguson Shawn , De Camilli Pietro
Published: May 2022

This method describes the usage of dextran-conjugated superparamagnetic iron nanoparticles (SPIONs) to purify a lysosome fraction from cultured cells.
eDNA extraction with kaneka v1
Authors: Xia Fei
Published: May 2022

We used the Kaneka Simple DNA Extraction Kit version 2 (Kaneka) for on-site DNA extraction from Sterivex. We injected 500µl of Solution A of the kit into the Sterivex and shook it by hand for 1min. We collected the Solution A buffer in the Sterivex by connecting the syringe with the connecting tube. We added 70µl of Solution B into the collected Solution A buffer in a 1.5-ml microtube and centrifuged (~2,000g) with a portable centrifuge for 3min. We collected 200µl of the supernatant solution.
2022 GenomeTrakr Proficiency Testing exercise (PulseNet Harmonized) v4
Authors: Balkey Maria , Timme Ruth , Haendiges Julie
Published: May 2022

This SOP outlines guidelines on how to process the isolates for the 2022 GenomeTrakr (GT) Proficiency Testing exercise. This SOP is applicable to all GenomeTrakr labs participating in the 2022 GenomeTrakr Proficiency Testing exercise (PulseNet Harmonized). The FDA GenomeTrakr program will ship the following proficiency testing isolates on April, 2022. ESP22-5191 Shigella SAP22-0236 Salmonella SAP22-0692 Salmonella SAP22-1300 Salmonella Completion of the entire proficiency test entails the following: GT participating laboratory generates sequencing data (fastq files) using the PT strains provided by CDC through FDA-CFSAN-WGS Program-GT in accordance to GT and/or PulseNet SOPs. Populate sample sheet according to 2022 GT Proficiency Testing exercise (PulseNet Harmonized) SOP Submission of sequencing records to the appropriate project on BaseSpace or Isilon according to GT SOPs By participating in the 2022 GT Proficiency Testing exercise (PulseNet Harmonized), GT labs provide consent to use the PT exercise data in subsequent analysis and manuscript publications. Participants will be acknowledged for their contribution on any publication that might require processing data from the 2022 GT PT exercise. v4: Added the file for GalaxyTrakr_PT_exercise_report_2022.xlsx
Production of a 1.8 billion CFU/ml yogurt using local products in Qatar v1
Authors: Jazi Al Naimi Al , Bendriss Ghizlane , Esteban Christina , Alansari Abdulaziz , Altamimi Alyaa , Al-Marri Maha , Ayoub Sama , Alnaimi Noor , Ibrahim Shahad , Manyama Jane , Gaskell Christine , Sebah Majda , McVeigh Clare
Published: May 2022

This protocol has been optimised to significantly improve the number of colony forming unit in local commercial yogurts. This protocol is an intervention on the starter Baladna (Qatar). This protocol increases its CFU/ml and resistance to stomach acidity. The end product has shown to contain more than 180 billion CFU in a jar of 100 ml, and highest resistance to acidity as compared to other yogurts tested.
transfer protocol 2 v1
Authors: not provided richard
Published: May 2022

test
transfer 4 v1
Authors: not provided richard
Published: May 2022

test
QSAR Prediction v1
Authors: Sciences BOC
Published: May 2022

The early linear free-energy approaches developed by Hansch and Free-Wilson have provided a fundamental scientific framework for the quantitative correlation of chemical structure with biological activity and spurred many developments in the field of quantitative structure-activity relationships (QSAR). QSAR prediction methods are attempting to predict the toxicological endpoint and breadth of mechanisms which are complicated. The quality and quantity of the available biological toxicology data could be another obstacle in the modeling process.
Preparation of Biological Tissues for Serial Block Face Scanning Electron Microscopy (SBEM) v2
Authors: J. Deerinck Thomas , A. Bushong Eric , H. Ellisman Mark , Thor Andrea
Published: May 2022

This protocol was designed to enhance signal for backscatter electron imaging of epoxy embedded mammalian tissue at low accelerating voltages (1-3 keV). However, it can easily be adapted for use with tissues from other species, tissue culture cells, plants and microbial cells by adjusting the buffer strength and the duration of relevant steps. This combinatorial heavy metal staining protocol employs a battery of contrasting steps after primary aldehyde fixation including: ferrocyanide reduced osmium tetroxide post fixation, thiocarbohydrazide-osmium liganding (OTO) and subsequent uranyl acetate and en bloc lead aspartate staining. Calcium chloride is included in a number of steps to enhance membrane preservation and staining. This protocol was designed primarily to emphasize the contrast of membranes. Many other contrasting agents may be included to increase staining of other cellular and extracellular constituents.
Computer-Aided Drug Discovery v1
Authors: Sciences BOC
Published: May 2022

Computer-aided drug design (CADD) is a widely used technology using computational tools and resources for the storage, management, analysis and modeling of compounds. It relies on digital repositories for the study of designing compounds with physicochemical characteristics, predicting whether a given molecule will be combined with the target, and if so how strongly. Computer-based methods can help us to search new hits in drug discovery, screening many irrelevant compounds at the same time and study the structure-activity relationship of drug molecules.
Wastewater QC workflow in GalaxyTrakr (SSQuAWK3) v5
Authors: Amirzadegan Jasmine , Kayikcioglu Tunc , not provided hugh.rand , Timme Ruth , Balkey Maria
Published: May 2022

PURPOSE: Step-by-step instructions for checking sequence quality for SARS-CoV-2 wastewater samples using SSQuAWK3: SARS - CoV - 2 Sequence Quality Assurance Workflow and Kontraption, version 3. The SSQuAWK3 workflow, implemented in CFSAN's custom Galaxy instance (GalaxyTrakr) will produce quality assessments for raw reads (Illumina MiSeq paired-end fastq files). SCOPE: This protocol covers the following tasks: 1. Set up an account in GalaxyTrakr 2. Create a new history 3. Upload data and reference files 4. Execute the SSQuAWK3 workflow 5. Interpret the results Protocol and SSQuAWK workflow version history: Protocol V1, SSQuAWK version 1: Basic protocol steps with screenshots Protocol V2, SSQuAWK version 1: Addition of a detailed 12 minute video tutorial Protocol V3, SSQuAWK version 2: Addition of 5 new genome mapping metrics Protocol V4 SSQuAWK version 3: Metrics now reported with fewer softwares, fewer underlying GalaxyTrakr jobs,and about 50% fewerunderlying GalaxyTrakr steps. Cleaner output table formats now include QC placeholder columns for SRA metadata template. Protocol V5 SSQuAWK version 3: Previous protocol version had broken links for FASTA and BED files, this version fixes the links.
Unclear insomnia types in randomized controlled trials and systematic reviews: the protocol for a meta-epidemiological study v2
Authors: Banno Masahiro , Tsujimoto Yasushi , Kohmura Kunihiro , Dohi Eisuke , Taito Shunsuke , Someko Hidehiro , Kataoka Yuki
Published: May 2022

Objectives. To examine tendencies and characteristics of randomized controlled trials (RCTs) and systematic reviews (SRs) that mention insomnia in abstracts and cannot be distinguished between insomnia disorder and insomnia symptoms (insomnia types) from the abstract and what is the intent of the unclear insomnia types described in the abstracts. Methods. This study is a meta-epidemiological study. We will include RCT and SR articles that include the word “insomnia” in methods or results of structured abstracts, published after 2010. We will exclude RCTs and SRs, which had neither methods nor results in non-structured abstracts. We will search MEDLINE via PubMed with English language restrictions. We will extract the information about whether abstracts of RCTs and SRs describe insomnia type clearly or not, and the characteristics. We will also extract information about the diagnosis of insomnia disorder for RCT and SR articles with unclear insomnia types and which intent insomnia disorder as insomnia in abstracts. We will report the proportion of RCT and SR articles with unclear insomnia types. We will compare characteristics between RCT and SR articles with unclear insomnia types and those with clear insomnia types. We will examine the above possible characteristics associated with the unclear insomnia types using multivariable logistic regression. We will report what was the intent of the unclear insomnia types described in the abstracts. We will also report what and how much proportion of diagnostic criteria was used for insomnia disorder for RCT and SR articles with unclear insomnia types. Ethics & Dissemination. This study does not need ethical approval. We registered this study protocol. We will publish the findings in a peer-reviewed journal and may present them at conferences. Discussion. This study will clarify what proportions and characteristics that do not describe insomnia type clearly in abstracts of RCTs and SRs. The results will inform the researchers, reviewers, and readers of the points that need special attention. Registration: Protocols.io
Nuclei isolation from snap-frozen human placental tissue for bulk ATACseq v1
Authors: Lindsay-Hewett Scott
Published: May 2022

This protocol describes the isolation of nuclei from snap-frozen human placental tissue for bulk ATACseq. It is a modified version of a protocol (SCBL Protocols - 10x Multiome Nuclei Isolation) supplied by the Jackson Laboratory (JAX), which is itself modified from Sigma Aldrich's Nuclei Isolation Kit: Nuclei EZ Prep protocol, and 10x Genomics' Demonstrated Protocol – Nuclei Isolation for Single Cell Multiome ATAC + Gene Expression Sequencing. It is recommended to work with no more than four samples at a time so that all samples can be processed in a timely manner.
Processing human frontal cortex brain tissue for population-scale Oxford Nanopore long-read DNA sequencing SOP v2
Authors: J Billingsley Kimberley , Dewan Ramita , Malik Laksh , Alvarez Jerez Pilar , Kiley Stith , Blauwendraat Cornelis , behalf of the CARD Long-read Team on
Published: May 2022

Processing human frontal cortex brain tissue for population-scale Oxford Nanopore long-read DNA sequencing SOP At the NIH's Center for Alzheimer's and Related Dementias (CARD) https://card.nih.gov/research-programs/long-read-sequencing we will generate long-read sequencing data from roughly 4000 patients with Alzheimer's disease, frontotemporal dementia, Lewy body dementia, and healthy subjects. With this research, we will build a public resource consisting of long-read genome sequencing data from a large number of confirmed people with Alzheimer's disease and related dementias and healthy individuals. To generate this large-scale nanopore sequencing data we have developed a protocol for processing and long-read sequencing human frontal cortex brain tissue, targeting an N50 of ~30kb and ~30X coverage. †Correspondence to: Kimberley Billingsley billingsleykj@nih.gov and Cornelis Blauwendraat cornelis.blauwendraat@nih.gov Acknowledgements: We would like to thank the Nanopore team (Androo Markham &Hannah Lucio), Circulomics Inc team (Jeffrey Burke, Michelle Kim, Duncan Kilburn & Kelvin Liu) and the whole CARD long-read team listed below => UCSC: Benedict Paten, Mikhail Kolmogorov, Miten Jain, Kishwar Shafin, Trevor Pesout; NHGRI: Adam Phillippy, Arang Rhie; Baylor: Fritz Sedlazeck; JHU: Winston Timp; NINDS: Sonja Scholz; NIA: Cornelis Blauwendraat, Kimberley Billingsley, Frank Grenn, Pilar Alvarez Jerez, Bryan Traynor, Shannon Ballard, Caroline Pantazis; CZI: Paolo Carnevali.
Tagmentation and library generation for human placental bulk ATACseq v1
Authors: Lindsay-Hewett Scott
Published: May 2022

This protocol describes the generation and amplification/indexing of tagmented libraries from freshly isolated nuclei for bulk ATACseq. It is adapted from Buenrostro et al., 2015, PMID: 25559105.
Nissl (Cresyl Violet) staining v1
Authors: not provided Christiana.bjorkli , Hovde Karoline , Monterotti Bruno
Published: May 2022

Nissl (Cresyl Violet) staining protocol
Plant assemble - Plant de novo genome assembly: annotation v2
Authors: Ferguson Scott , Jones Ashley , Borevitz Justin
Published: May 2022

With the advancement of long-read sequencing technologies and associated bioinformatics tools, it has now become possible to de novo assemble complex plant genomes with unrivalled contiguity, completeness and correctness. As read lengths can surpass repeat lengths, the ability to assemble genomes de novo has dramatically improved, whereby complex plant genomes of widely variable sizes and repeat content have highly benefited. Despite these improvements, challenges remain in performing de novo assembly, namely in developing a reliable workflow and in tool choice. Here we present a protocol collection of bioinformatic workflows detailing plant genome assembly using Oxford Nanopore Technologies long-reads with a de novo assembler (Canu), syntenic or Hi-C scaffolding, and RNA and/or gene homology-based annotation. We have developed and tested these protocols on multiple plant genomes. Using these protocols with sufficient coverage of long-reads, a highly contiguous, complete, and correct plant genome can be assembled. These genomes can further genomic research into structural variation among groups, and SNP genotyping and association studies among populations.
Plant assemble - Plant de novo genome assembly: quality assessment v2
Authors: Ferguson Scott , Jones Ashley , Borevitz Justin
Published: May 2022

With the advancement of long-read sequencing technologies and associated bioinformatics tools, it has now become possible to de novo assemble complex plant genomes with unrivalled contiguity, completeness and correctness. As read lengths can surpass repeat lengths, the ability to assemble genomes de novo has dramatically improved, whereby complex plant genomes of widely variable sizes and repeat content have highly benefited. Despite these improvements, challenges remain in performing de novo assembly, namely in developing a reliable workflow and in tool choice. Here we present a protocol collection of bioinformatic workflows detailing plant genome assembly using Oxford Nanopore Technologies long-reads with a de novo assembler (Canu), syntenic or Hi-C scaffolding, and RNA and/or gene homology-based annotation. We have developed and tested these protocols on multiple plant genomes. Using these protocols with sufficient coverage of long-reads, a highly contiguous, complete, and correct plant genome can be assembled. These genomes can further genomic research into structural variation among groups, and SNP genotyping and association studies among populations.
Plant assemble - Plant de novo genome assembly: assembly v2
Authors: Ferguson Scott , Jones Ashley , Borevitz Justin
Published: May 2022

With the advancement of long-read sequencing technologies and associated bioinformatics tools, it has now become possible to de novo assemble complex plant genomes with unrivalled contiguity, completeness and correctness. As read lengths can surpass repeat lengths, the ability to assemble genomes de novo has dramatically improved, whereby complex plant genomes of widely variable sizes and repeat content have highly benefited. Despite these improvements, challenges remain in performing de novo assembly, namely in developing a reliable workflow and in tool choice. Here we present a protocol collection of bioinformatic workflows detailing plant genome assembly using Oxford Nanopore Technologies long-reads with a de novo assembler (Canu), syntenic or Hi-C scaffolding, and RNA and/or gene homology-based annotation. We have developed and tested these protocols on multiple plant genomes. Using these protocols with sufficient coverage of long-reads, a highly contiguous, complete, and correct plant genome can be assembled. These genomes can further genomic research into structural variation among groups, and SNP genotyping and association studies among populations.
Purifying Vps13-VAB domain proteins v1
Authors: not provided jyoti.adlakha
Published: May 2022

In this protocol, we have listed the steps to purify the Vps13 adaptor binding domain (or VAB) with or without PxP motif fusion peptide derived from adaptor proteins Mcp1, Ypt35 or Spo71.
Crystallization, data collection and structure solution of PxP-VAB v1
Authors: not provided jyoti.adlakha
Published: May 2022

Protocol lists the steps to obtain PxP(Mcp1ct)-VABct crystals, X-ray diffraction data acquisition and obtaining the structure solution using molecular replacement with a model generated by AlphaFold2.
FLASH-seq protocol v3
Authors: Picelli Simone , Hahaut Vincent
Published: May 2022

The single-cell RNA-sequencing (scRNA-seq) field has evolved tremendously since the first paper was published back in 2009. While the first methods analysed just a handful of cells, the throughput and performance rapidly increased over a very short timespan. However, it was not until the introduction of emulsion droplets methods, that the robust and reproducible analysis of thousands of cells became feasible. Despite generating data at a speed and a cost per cell that remains unmatched by full-length protocols like Smart-seq, scRNA-seq in droplets still comes with the drawback of addressing only the terminal portion of the transcripts, thus lacking the required sensitivity for comprehensively analyzing the transcriptome of individual cells. Building upon the existing Smart-seq2/3 workflows, we developed FLASH-seq (FS), a new full-length scRNA-seq method capable of detecting a significantly higher number of genes than both previous versions, requiring limited hands-on time and with a great potential for customization.
Single-Cell Isolation of Human Articular Cartilage v1
Authors: not provided hswahn , Lotz Martin
Published: May 2022

This is a protocol that describes the process of isolating single cells from healthy knee cartilage for single cell RNAseq.
Quantification of the SARS-CoV-2 using electronegative membrane filtration and dPCR v2
Authors: Narum Solana , Stalder Thibault , Coats Erik , Top Eva
Published: May 2022

This protocol was developed in an effort to serve as a timely and efficient method for the surveillance of the SARS-CoV-2 in primary influent wastewater samples. This process describes viral concentration via electronegative adsorption with pH adjustment, RNA extraction using the MP Biomedicals FastPrep-24 and the Qiagen AllPrep PowerViral DNA/RNA kits with the Qiacube Connect, and quantification of the N1 and N2 genes in SARS-CoV-2 using the GT Digital SARS-CoV-2 Wastewater Surveillance for QIAcuity. To compile the entire process from beginning to the end some sections were taken from the AllPrep PowerViral DNA/RNA Kit (Qiagen) and the GT Digital SARSCoV- 2 Wastewater Surveillance for QIAcuity v1.0 (GT Molecular) handbooks.
SAPAP3 Genotyping Protocol v1
Authors: not provided kajsbr
Published: May 2022

Protocol for SAPAP3 KO Mouse Model Genotyping from Dr. Lisa Gunaydin's Lab, University of California, San Francisco
Unclear insomnia types in randomized controlled trials and systematic reviews: the protocol for a meta-epidemiological study v3
Authors: Banno Masahiro , Tsujimoto Yasushi , Kohmura Kunihiro , Dohi Eisuke , Taito Shunsuke , Someko Hidehiro , Kataoka Yuki
Published: May 2022

Objectives. To examine tendencies and characteristics of randomized controlled trials (RCTs) and systematic reviews (SRs) that mention insomnia in abstracts and cannot be distinguished between insomnia disorder and insomnia symptoms (insomnia types) from the abstract and what is the intent of the unclear insomnia types described in the abstracts. Methods. This study is a meta-epidemiological study. We will include RCT and SR articles that include the word “insomnia” in methods or results of structured abstracts, published after 2010. We will exclude RCTs and SRs, which had neither methods nor results in non-structured abstracts. We will search MEDLINE via PubMed with English language restrictions. We will extract the information about whether abstracts of RCTs and SRs describe insomnia type clearly or not, and the characteristics. We will also extract information about the diagnosis of insomnia disorder for RCT and SR articles with unclear insomnia types and which intent insomnia disorder as insomnia in abstracts. We will report the proportion of RCT and SR articles with unclear insomnia types. We will compare characteristics between RCT and SR articles with unclear insomnia types and those with clear insomnia types. We will examine the above possible characteristics associated with the unclear insomnia types using multivariable logistic regression. We will report what was the intent of the unclear insomnia types described in the abstracts. We will also report what and how much proportion of diagnostic criteria was used for insomnia disorder for RCT and SR articles with unclear insomnia types. Ethics & Dissemination. This study does not need ethical approval. We registered this study protocol. We will publish the findings in a peer-reviewed journal and may present them at conferences. Discussion. This study will clarify what proportions and characteristics that do not describe insomnia type clearly in abstracts of RCTs and SRs. The results will inform the researchers, reviewers, and readers of the points that need special attention. Registration: Protocols.io
A real-time PCR method to genotype mutant mouse models with altered affinity for cardiotonic steroids on the Na,K-ATPase v2
Authors: W W Chomczynski Peter , M Vires Kianna , Rymascewski Michal , A. Heiny Judith
Published: May 2022

The highly conserved, cardiotonic steroid binding site (also termed ouabain binding site) on the primary α subunit of Na,K-ATPase plays a receptor signaling role in a range of vital cell processes and is a therapeutic target for human disease. Mouse lines with altered affinity for cardiotonic steroids on the α1 or α2 subunit isoform of Na,K-ATPase, developed by the late Dr. Jerry Lingrel and collaborators, are a valuable tool for studying its physiological roles and drug actions.In one model, the normally ouabain resistant α1 isoform was rendered sensitive to ouabain binding. In a second model, the normally sensitive α2 isoform was rendered resistant to ouabain binding.Additional useful models are obtained by mating these mice. To further advance their use, we developed a rapid, real-time PCR method that amplifies digested tail clip samples without the need to isolate DNA, and detects mutant alleles using specific primers and fluorescent probes. PCR is performed in fast mode with up to 15 samples processed in 40 min. The method was validated by comparing results with a previous two-step method using PCR amplification followed by gel electrophoresis, and by Sanger sequencing. We also clarified inconsistencies in published sequences, updated numbering to current reference sequences, and confirmed the continued presence of the mutations in the colony. It is expected that the wider availability of these models and a more efficient genotyping protocol will advance studies of the Na,K-ATPase and its cardiotonic steroid receptor.