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Authors: Grant Mccallum, Nathan Kostick, Joseph Marmerstein, Yang Zheng, Dominique Durand
Group: SPARC
Summary: We propose to develop a neural interface based on carbon nanotube (CNT) yarns with a size similar to large axons (10 µm) that has a mechanical and molecular compatibility with the nerve. The mechanical properties of the interface will be optimized by matching flexural rigidity of the implant to that of the glossopharyngeal nerve (GLN). Since the wire is very flexible, a custom-inserting technique has been developed for this purpose and tested in rats.  Following insertion, the signal-to-noise ratio and impedance of the wires will be measured for 3 months. The biocompatibility of the implant will be assessed using histological techniques.

Proper citation: Grant Mccallum, Nathan Kostick, Joseph Marmerstein, Yang Zheng, Dominique Durand 2021. Glossopharyngeal Nerve Chronic Recording In Anesthetized Rat. protocols.io https://dx.doi.org/10.17504/protocols.io.wssfeee Copy   

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Authors: Yuki Kataoka, Shiho Oide, Takashi Ariie, Yasushi Tsujimoto, Toshi A. Furukawa
Summary: Background:No studies have comprehensively assessed the quality of studies related to COVID-19 in preprints [11]. Hence, we will conduct a meta-epidemiological study based on the following research question: Are the quality of COVID-19 RCT and SR articles in medRxiv lower than those indexed in PubMed? Methods:We will conduct a meta-epidemiological cross-sectional study. We will include randomized controlled trial articles and systematic review articles indexed in PubMed or MedRxiv from 1st January to 15th June 2020. We will include articles of COVID-19 patients or healthcare workers engaged in the care of COVID-19 patients. We will exclude study protocols. We will evaluate the characteristics of included studies, methodological quality, number of mentions on social networking sites (SNS), and citations. We will use chi-squared test, logistic regression analysis, and linear regression analysis as appropriate. Two-tailed p values will be considered statistically significant if less than 0.05. Ethical consideration:We will only use openly available data. There is no need to make ethical considerations.

Proper citation: Yuki Kataoka, Shiho Oide, Takashi Ariie, Yasushi Tsujimoto, Toshi A. Furukawa 2020. Quality of COVID-19 research in preprints: a meta-epidemiological study protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.bhm8j49w Copy   

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Authors: Cristian Riccio

Proper citation: Cristian Riccio 2019. Defrost a C. elegans strain from the -80 C freezer. protocols.io https://dx.doi.org/10.17504/protocols.io.5xug7nw Copy   

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Authors: Yasmin Bar El
Group: GigaScience Press
Summary: Primary cortical neuronal-astrocyte cell culture preparation. Dissociation, preparation and plating of mice cortex neurons and glia cells on MEA.

Proper citation: Yasmin Bar El 2018. Neuron-astrocyte culture preparation. protocols.io https://dx.doi.org/10.17504/protocols.io.tqaemse Copy   

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Authors: Carine Puppo, Tomas Voisin, Brigitte Gontero
Group: High molecular weight DNA extraction from all kingdoms
Summary: Asterionella formosa is a freshwater pennate diatom that forms star-shaped colonies, whose mitochondrial DNA was recently sequenced [1]. This diatom is of interest for genetic studies [2,3] and for the study of photosynthesis [4,5]. However, extraction of genomic DNA (gDNA) from A. formosa is difficult because of its silica cell-wall and the presence of photosynthetic pigments that contaminate the DNA. Here, we present an optimised protocol for gDNA extraction from A. formosa that overcomes those two problems. We also assessed its efficacity in yielding high-purity gDNA compared to a standard protocol using hexadecyltrimethylammonium bromide (CTAB).References[1] Villain A. et al. (2017) Mitochondr. DNA. 2 (1), 97-98.[2] De Bruin A. et al. (2004) J. Phycol. 40, 823-830.[3] van Den Wyngaert S. et al. (2015) Mol. Ecol. 24, 2955-2972.[4] Boggetto N. et al. (2007) J. Phycol. 43, 1227-1235.[5] Erales J. et al. (2008) J. Phycol. 44, 1455-1464.

Proper citation: Carine Puppo, Tomas Voisin, Brigitte Gontero 2017. Genomic DNA extraction from the pennate diatom Asterionella formosa optimised for next generation sequencing.. protocols.io https://dx.doi.org/10.17504/protocols.io.jytcpwn Copy   

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Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This protocol describes the system that is used to capture Z stack images which are reassembled post-processing into 3D reconstructions.Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Proper citation: Allen Institute for Brain Science 2020. Bright Field Image Capture with Zeiss AxioImager. protocols.io https://dx.doi.org/10.17504/protocols.io.bdpmi5k6 Copy   

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Authors: Eftychis Frangedakis, Marta Tomaselli, Marius Rebmann, Susana Sauret-Gueto
Group: OpenPlant Project
Summary: This protocol allows for quick and dirty genomic DNA extraction. It can easily be used for genotyping with PCR. The quality of the genomic DNA extracted is not suitable for any other application.

Proper citation: Eftychis Frangedakis, Marta Tomaselli, Marius Rebmann, Susana Sauret-Gueto 2019. Marchantia genotyping (quick and dirty genomic DNA extraction). protocols.io https://dx.doi.org/10.17504/protocols.io.4wagxae Copy   

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Authors: Kristoffer Bach Falkenberg, Cristina Hernandez Rollan, Maja Rennig, Andreas Birk Bertelsen, Morten Norholm
Summary: B. subtilis is a gram-positive bacteria used by both academia and industry as a protein production workhorse. This is due to its' their excellent fermentation properties, high production titers, and capacity to secrete proteins into the extracellular medium.This protocol describes how to express proteins in B. subtilis. The protocol is developed using KO7-S, although it might also work for other strains as well. The method is adapted from Rasmussen, M. D.; Bjoernvad, M. E.; Diers, I. Pectate Lyase Fusion for Expression and Secretion of Polypeptides. WO 00/75344, 2000 and Jensen, K.; Østergaard, P. R.; Wilting, R.; Lassen, S. F. Identification and Characterization of a Bacterial Glutamic Peptidase. BMC Biochem.2010, 11 (1), 47. https://doi.org/10.1186/1471-2091-11-47.

Proper citation: Kristoffer Bach Falkenberg, Cristina Hernandez Rollan, Maja Rennig, Andreas Birk Bertelsen, Morten Norholm 2020. Protein expression in Bacillus subtilis. protocols.io https://dx.doi.org/10.17504/protocols.io.bdmui46w Copy   

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Authors: Wei-Ting Lu, Ming Jiang, Robert Goldstone, Karen Ambrose, Chris Ekin, Amy Strange, Nnenna Kanu, Paul Grant, Efthymios Fidanis, Jerome Nicod, David Moore, Michael Howell
Group: BGI, Coronavirus Method Development Community, Crick COVID-19 Consortium
Summary: Purpose of examination / Clinical relevanceAt the end of 2019, several pneumonia cases were reported in Wuhan, China and the pathogen was confirmed as a new viral strain. World Health organization has named the newly identified coronavirus as 2019-nCoV, also known as COVID19. The disease developed into a dangerous pandemic, posing major challenges to the NHS. Although more research is necessary to better understand the virus, in response to the emergency, simple and rapid testing is essential to identify the virus in infected individuals. This will aid the implementation of efficient interventions to contain the spread, and distinguish healthcare workers who have been infected, and are required to self-isolate, from those showing similar symptoms but which are not 2019-nCoV associated. The latter category may continue to work, alleviating stress on hard-pressed healthcare resources. 2019-nCoV is an RNA virus, and the diagnostic tests detect viral RNA in swabs from patient airways using a reverse transcriptase PCR assay. Samples are submitted to HSL, an accredited reporting laboratory, and transferred to the Crick for testing. The first step of the process is sample receipt at the Crick.This protocol describes the preparation of the RT-PCR master mix plate, automated addition of RNA template and the reverse-transcriptase quantitative PCR of viral RNA to detect the Orf1ab region of the viral genome, which has high specificity to the 2019-nCoV virus. Furthermore, the protocol describes the analysis of the COVID-19 test results.Principles of ExaminationThe kit is a qualitative in vitro nucleic acid amplification assay to detect the new coronavirus 2019-nCoV identified in China in 2019 using Reverse transcription PCR of RNA derived from specimen of throat swabs and Bronchoalveolar Lavage Fluid (BALF) from individuals suspected to have been exposed to the virus. The kit is based on in vitro RT-PCR with fluorescent probes. Primers and sequence-specific fluorescence probes were designed tailored to the high conservative ORF1ab region of the 2019-nCoV genome. The probes are oligonucleotides with attached fluorophores at the 5' end (FAM as reporter) and at the 3’ end (quencher). In addition, a specific primer and probe set designed to human b-actin is included as an internal reference with fluorophores VIC/HEX attached at 5’ end as reporter. During the PCR procedure, the DNA polymerase cleaves the probe at the 5’ end and separates the reporter dye from the quencher dye when the probes hybridize to the target DNA. This cleavage results in a fluorescent signal generated by the cleaved reporter dye, which is monitored in real-time by the PCR detection system. Monitoring the fluorescence intensities during Real Time allows the qualitative detection of 2019-nCoV in specimens.Analysis of the COVID19 test results will allow the qualitative detection of the 2019-nCoV nucleic acid in specimens from patients with suspected 2019n-nCoV. This process will enable the assessment of the infection in clinical and public health practice. The workflow for reporting results from this test is illustrated below:

Proper citation: Wei-Ting Lu, Ming Jiang, Robert Goldstone, Karen Ambrose, Chris Ekin, Amy Strange, Nnenna Kanu, Paul Grant, Efthymios Fidanis, Jerome Nicod, David Moore, Michael Howell 2020. SARS-CoV-2 detection using BGI RT-PCR kit. protocols.io https://dx.doi.org/10.17504/protocols.io.bfc8jizw Copy   

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Authors: Jiaqi Liang
Summary: In this paper, we apply statistics and network analysis methods to explore the dynamic characteristics of three transaction networks. We download transaction data from the respective blockchain explorers. To the best of our knowledge, these are the largest datasets adopted in cryptocurrency analysis to date. We analyze the growth pattern of the accumulated network and find that unlike most networks, these cryptocurrency networks do not always densify over time. Then based on the datasets, we find that the monthly repetition ratios measured by either node or edge are relatively low. As such, studying the whole accumulated network, as done in most previous work, is not the appropriate way to understand the network dynamics. Hence we focus on coining the dynamics through computing the values of typical network measures on a monthly basis, and make a comparison among the three networks. 

Proper citation: Jiaqi Liang 2018. Evolutionary Dynamics of Cryptocurrency Transaction Networks. protocols.io https://dx.doi.org/10.17504/protocols.io.rc3d2yn Copy   

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Authors: Mengchen Shi

Proper citation: Mengchen Shi 2017. Analysis of apoptosis by AnnexinPE/7-aminoactinomycin D (7-AAD) staining. protocols.io https://dx.doi.org/10.17504/protocols.io.kcecste Copy   

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Authors: Camila Rangel Smith, Kasra Hosseini
Group: Vivarium Population Spenser
Summary: Description of the steps followed by Vivarium Population Spenser library when running the Internal migration module.

Proper citation: Camila Rangel Smith, Kasra Hosseini 2020. Vivarium Population Spenser: Internal migration module. protocols.io https://dx.doi.org/10.17504/protocols.io.bn9imh4e Copy   

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Authors: Frank Brosius
Group: Diabetic Complications Consortium
Summary: This protocol describes the procedures for quantifying the percent area of the glomerulus that is PAS stained and is expressed as the mesangial index.Diabetic Complication:

Proper citation: Frank Brosius 2019. Mesangial Index Quantification. protocols.io https://dx.doi.org/10.17504/protocols.io.8e7hthn Copy   

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Authors: Roey Angel, Eva Petrova
Group: SoWa RI Anaerobic and Molecular Microbiology (public)
Summary: This protocol describes how to quantify 16S rRNA bacterial gene or transcript copy numbers using Droplet Digital PCR technology (ddPCR) from Bio-Rad This is an up-to-date modification of a classical bacterial enumeration qPCR-assay. This assay uses the EvaGreen™ chemistry. The primers are taken from Yu et al. (2005).

Proper citation: Roey Angel, Eva Petrova 2020. Quantification of 16S rRNA Gene Copies Using ddPCR (EvaGreen-based assay: 338F-805R). protocols.io https://dx.doi.org/10.17504/protocols.io.bmqwk5xe Copy   

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Authors: Sam Li
Group: BioLegend

Proper citation: Sam Li 2019. Immunohistochemistry Protocol for Ultra Streptavidin Detection Kits (USA). protocols.io https://dx.doi.org/10.17504/protocols.io.95rh856 Copy   

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Authors: Carlos Helbig
Summary: This protocol describes how to transform chmically competent Vibrio natriegens cells.The protocol was described and published by Weinstock et al., 2016

Proper citation: Carlos Helbig 2018. Transformation of Vibrio natriegens. protocols.io https://dx.doi.org/10.17504/protocols.io.psudnew Copy   

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Authors: Renuka Kudva, Andreas Vogt, Kärt Denks, Gunnar von Heijne
Summary: Isolation of ribosome-nascent chains of co-translationally folded domains. Detailed description of protocol published in https://doi.org/10.1073/pnas.1810523115. The arrest sequence used for stalling was TnaC (the leader peptide of the tryptophanase operon).

Proper citation: Renuka Kudva, Andreas Vogt, Kärt Denks, Gunnar von Heijne 2019. Isolation of ribosome-associated nascent chains of soluble proteins produced in Escherichia coli. protocols.io https://dx.doi.org/10.17504/protocols.io.23fggjn Copy   

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Authors: Joao Vitor Molino
Summary: X7 is a a Pfu-sso7d fusion DNA polymerase that is compatible with uracil-excision cloning and has been used with sucess to amplify our high GC-content sequences and it is a valuable tool for USER clonning.Before startingPlease note that protocols with X7 may differ from protocols with other polymerases. Conditions recommended below were tested with our sequences, mostly high GC-content sequences from Chlamydomonas reinhardtii expression vectors. Please refer to the original work that develop the fusion polymerase. All components should be mixed prior to use. Reference:Nørholm, M. H. H. (2010). A mutant Pfu DNA polymerase designed for advanced uracil-excision DNA engineering. BMC Biotechnology, 10. https://doi.org/10.1186/1472-6750-10-21

Proper citation: Joao Vitor Molino 2020. High GC-content PCR. protocols.io https://dx.doi.org/10.17504/protocols.io.bprimm4e Copy   

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Authors: Christian Lawrence, Jason Best, Althea James, Shane Hurley, Mitchel Shia, Michelle Urh
Summary: Environmental and housing conditions experienced by laboratory animals exert profound effects on their biology, physiology, and behavior.  These parameters are important and often overlooked sources of potential variation in experiments, and should be reported in peer-reviewed publications in order to promote scientific reproducibility.  To that end, here we provide a summary of the environmental conditions in zebrafish (Danio rerio) aquaculture facilities at Boston Children's Hospital (BCH).   We include data on the physico-chemical, health, and nutrition of zebrafish in three separate facilities at BCH: Karp, Enders SSB, and Enders Lobby*. In this year's version, we also include new information on our sanitization protocols and equipment. We also provide some additional characterization of our sampling methods for water parameters and health in a legend. This information will be applicable to any study involving zebrafish conducted in one of these three facilities during the year of 2018.*version amended to include new information on diet in this facility.

Proper citation: Christian Lawrence, Jason Best, Althea James, Shane Hurley, Mitchel Shia, Michelle Urh 2019. Zebrafish (Danio rerio) Environmental Summary, Aquatic Resources Program, Boston Children's Hospital 2018. protocols.io https://dx.doi.org/10.17504/protocols.io.w36fgre Copy   

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Authors: 宏亮 董
Group: 2019 iGEM NEFU_China
Summary: This protocol includes three different methods of digestion.

Proper citation: 宏亮 董 2019. Restriction Digest. protocols.io https://dx.doi.org/10.17504/protocols.io.7h6hj9e Copy   

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