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Authors: Hannah Gunderman
Summary: This protocol is a description of how Hannah Gunderman facilitates her focus groups around Doctor Who episodes in order to better understand how viewers perceive messages of empathy, geography, and social themes in the episodes. In this example, Hannah describes her focus group protocol for the Doctor Who episode "Vincent and the Doctor" (Series 5, episode 10). This protocol is meant to demonstrate how the protocols.io platform can be used to document workflows and methods within humanities researcher, showing how the platform can be for any researcher regardless of domain area.

Proper citation: Hannah Gunderman 2020. Focus group methodology for Doctor Who viewing behaviors  . protocols.io https://dx.doi.org/10.17504/protocols.io.bjjykkpw Copy   

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Authors: Maximilian Krause
Group: The Nordic Nanopore-Seq Network
Summary: Oxford Nanopore Technologies allows sequencing of native RNA for the first time. Additionally they released tiny devices that democratize sequencing among scientists. However, the smallest sequencing device - Flongle - is so far not officially supported for RNA sequencing experiments.This protocol provides users with a personally tested framework protocol to use Flongle flow-cells for native RNA sequencing. It is based on the protocol for ONT RNA library preparation and changed to fit the volume and input requirements for Flongle.The here advised workflow will routinely result in 100,000 native RNA reads within 24 hours of sequencing.

Proper citation: Maximilian Krause 2020. Flongle DirectRNA Library preparation. protocols.io https://dx.doi.org/10.17504/protocols.io.bcwcixaw Copy   

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Authors: Tonis Org, Kati Hensen, Rita Kreevan, Elina Mark, Olav Sarv, Reidar Andreson, Ülle Jaakma, Andres Salumets, Ants Kurg
Summary: Chromatin immunoprecipitation coupled with next-generation sequencing (ChIP-seq) has revolutionized our understanding of chromatin-related biological processes. The method, however, requires thousands of cells and has therefore limited applications in situations where cell numbers are limited. Here we describe a novel method called Restriction Assisted Tagmentation Chromatin Immunoprecipitation (RAT-ChIP) that enables global histone modification profiling from as few as 100 cells. The method is simple, cost-effective and takes a single day to complete.

Proper citation: Tonis Org, Kati Hensen, Rita Kreevan, Elina Mark, Olav Sarv, Reidar Andreson, Ülle Jaakma, Andres Salumets, Ants Kurg 2019. RAT-ChIP - Restriction Assisted Tagmentation Chromatin Immunoprecipitation . protocols.io https://dx.doi.org/10.17504/protocols.io.69qhh5w Copy   

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Authors: Sonia Hall

Proper citation: Sonia Hall 2015. Western Blot Sample Prep- Whole Drosophila Embryos. protocols.io https://dx.doi.org/10.17504/protocols.io.cnzvf5 Copy   

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Authors: Rachele Cesaroni
Group: Protist Research to Optimize Tools in Genetics (PROT-G)

Proper citation: Rachele Cesaroni 2017. Small-scale silencing experiment in vegetative Euplotes crassus (provisional). protocols.io https://dx.doi.org/10.17504/protocols.io.hi5b4g6 Copy   

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Authors: Rekha Seshadri
Group: ECOGEO
Summary: Introduction to functional annotation and Integrated Microbial Genomes (IMG) at the Joint Genome Institute (JGI).Open this protocol inside the virtual machine (details in 'Start Instructions') for easy copy, paste of commands into the command line terminal window. 

Proper citation: Rekha Seshadri 2016. ECOGEO 'Omics Training: 4.2 Annotation. protocols.io https://dx.doi.org/10.17504/protocols.io.fi8bkhw Copy   

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Authors: Anita Bandrowski

Proper citation: Anita Bandrowski 2020. this is a test. protocols.io https://dx.doi.org/10.17504/protocols.io.2g9gbz6 Copy   

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Authors: Dr. Steven Wilhelm
Group: The Aquatic Microbial Ecology Research Group - AMERG (The Buchan, Zinser and Wilhelm labs), CyanoHABs
Summary: Please contact Dr. Steven Wilhelm ([email protected]) for additional information regarding this protocol.

Proper citation: Dr. Steven Wilhelm 2017. BG-11 Trace metal mix A5. protocols.io https://dx.doi.org/10.17504/protocols.io.h92b98e Copy   

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Authors: Oliver Fiehn
Group: Mouse Metabolic Phenotyping Centers, Metabolomics Protocols & Workflows
Summary: Summary:This SOP describes sample extraction and sample preparation for lipid profiling by liquid chromatography / quadrupole time of flight mass spectrometry (LC-QTOF) or nanoelectrospray ion trap-FTICR MS.

Proper citation: Oliver Fiehn 2019. UC Davis - Metabolomics: Sample preparation for Lipidomics. protocols.io https://dx.doi.org/10.17504/protocols.io.ytpfwmn Copy   

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Authors: Igem Dusseldorf
Summary: Removal of genomic DNA from RNA preparations

Proper citation: Igem Dusseldorf 2019. Removal of gDNA from totalRNA. protocols.io https://dx.doi.org/10.17504/protocols.io.8bfhsjn Copy   

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Authors: Angel Justiz-Vaillant
Summary: This ELISA was used to study Staphylococcal protein-A (SpA) interactions with various mammalian and avian immunoglobulins.

Proper citation: Angel Justiz-Vaillant 2021. Protein A- Protein A Sandwich ELISA. protocols.io https://dx.doi.org/10.17504/protocols.io.bsu4neyw Copy   

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Authors: Sara Calvert
Summary: The main purpose of pregnancy testing as part of a clinical research protocol is to minimize risk to an embryo or fetus from exposure to potentially harmful study interventions.  The specific goal of testing may be to prevent exposure completely (by testing prior to any study interventions), or to minimize the duration of exposure during early pregnancy (by periodic testing during the study).This survey provided five clinical study scenarios and asked respondents a series of questions about the acceptable risk of an unintended fetal exposure to a drug, the importance of several factors going into clinical study design, and the type of pregnancy testing protocol that you would recommend.

Proper citation: Sara Calvert 2018. CTTI Pregnancy Testing in Clinical Trials Project Survey. protocols.io https://dx.doi.org/10.17504/protocols.io.rj6d4re Copy   

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Authors: Eugene Drokhlyansky, Nicholas Van Wittenberghe, Michal Slyper, Julia Waldman, Asa Segerstolpe, Orit Rozenblatt-Rosen, Aviv Regev
Group: NCIHTAN
Summary: This protocol describes a nuclei isolation method for frozen tissues based on the previously published protocol by Gao et al. and modified by Drokhlyansky et al. It can be used on both healthy and disease tissues and is compatible with droplet-based single-nucleus RNA-Seq technology Slyper et al.This method is part of a toolbox for processing frozen tissue samples for single-nucleus RNA-Seq, including the NST (this protocol), CST and TST protocols (all available in protocols.io). We recommend users to test all three protocols on their tissue type of interest and perform side-by-side comparison of the data generated.  A description of the complete toolbox and guidance for testing and selecting methods from the toolbox for processing other tumors can be found in Slyper et al.For the Human Tumor Atlas Pilot Project (HTAPP), this protocol was tested on metastatic breast cancer, ovarian cancer and pediatric neuroblastoma frozen samples. For these cancer types, however, TST was the protocol of choice. When tested on HTAPP samples, NST, in general, underperformed compared to TST and CST based on several QC parameters.

Proper citation: Eugene Drokhlyansky, Nicholas Van Wittenberghe, Michal Slyper, Julia Waldman, Asa Segerstolpe, Orit Rozenblatt-Rosen, Aviv Regev 2020. HTAPP_NST- Nuclei isolation from frozen tissue. protocols.io https://dx.doi.org/10.17504/protocols.io.bhbdj2i6 Copy   

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Authors: Judy Northill, Ian Mackay
Group: protocols.io Ambassadors, Public Health Virology, Forensic and Scientific Services
Summary: A real-time RT-PCR to detect the "novel Wuhan" betacoronavirus or Wuhan seafood market pneumonia virus. Based on sequence MN908947 made available by Professor Yong-Zhen Zhang, Fudan University, Shanghai, China. The target region is within the ORF1ab sequence.Not tested on wild-type virus (as of 25Jan2020), it is expected to be capable of only detecting the Wuhan coronavirus.NotesAssay is fully optimised (as of 24Jan2020).A final name for this virus has not been decided (as of 25Jan2020).

Proper citation: Judy Northill, Ian Mackay 2020. Wuhan coronavirus (2019-nCoV) real-time RT-PCR ORF1ab 2020 (Wuhan-ORF1ab). protocols.io https://dx.doi.org/10.17504/protocols.io.bbsginbw Copy   

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Authors: Bonnie Hurwitz
Group: Hurwitz Lab, ABE487_HMP, ABE487_EMP
Summary: This tutorial shows you how to create a pipeline on the University of Arizona HPC.  Specifically, we will be creating a pipeline to run blast, using scripts you are writing or have already written from ABE 487 weeks 10 and 11.  The pipeline will do the following:1.  split each of the files from the night/day transcriptome into files with 10,000 sequences each2.  run blastp against  uniprot_sprot.fasta3.  parse the blast results

Proper citation: Bonnie Hurwitz 2015. Building an HPC blast pipeline. protocols.io https://dx.doi.org/10.17504/protocols.io.d5a82d Copy   

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Authors: Luciano Martelotto
Summary: A1Please note these are just my notes and by no means are ground truth of how Multi-seq needs to be performed. I WELCOME to comments, suggestions and amendments.

Proper citation: Luciano Martelotto 2020. Multi-Seq: my notes from the lab. protocols.io https://dx.doi.org/10.17504/protocols.io.bejyjcpw Copy   

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Authors: Tina Lasisi
Summary: This protocol prepares hair fibers per sample for curvature analysis.Diagrams were created with BioRender.com

Proper citation: Tina Lasisi 2020. Sample preparation protocol for hair fiber curvature analysis. protocols.io https://dx.doi.org/10.17504/protocols.io.bbweipbe Copy   

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Authors: Marcos Viana, Mauricio Mudadu, Adhemar Zerlotini
Summary: A1The Plantannot software provides several filters and a text search box that allows searching for molecules by its desired annotation features. These filters are needed to obtain PUFs and to try to relate them to abiotic stresses using RNA-seq expression data and co-expression networks. The Filters menu is separated in 8 fields, of those we are going to use only five: “Organism”, “Feature type”, “Orthology”, “Orthologs_coexpression” and “Analyses”. The “Feature Type” filter has three molecule types, from those the polypeptide box is the only that is going to be always checked and the others blank. By using the other 4 remaining filters, 6 protocols were created as examples of different ways to selecting PUFs. Protocol A: using lack of both homology and protein domain signatures. Protocol B: using lack of homology, presence of domain signatures - trying to select Domains of Unknown Function (DUF) from PFAM, and the text search “Unknown function”.Protocol C: using homology, lack of protein domain signatures and the text search “Unknown function”. Protocol D-F: same protocols of A-C but using ortholog groups to find homolog proteins with co-expression data related to abiotic stress.2Protocol C is intended do find PUFs from organisms that proteins are already public in the NCBI´s nr database and have no protein domains found by Interproscan. Proteins will be selected using the text search "Unknown function".

Proper citation: Marcos Viana, Mauricio Mudadu, Adhemar Zerlotini 2020. Find Proteins of Unknown Function (PUFs) using Plantannot - Protocol C. protocols.io https://dx.doi.org/10.17504/protocols.io.bgdijs4e Copy   

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Authors: Matthew Sullivan
Group: VERVE Net, Sullivan Lab
Summary: For One-step growth curves for Cellulophaga phages protocol.

Proper citation: Matthew Sullivan 2016. Centrifuged Sample Steps. protocols.io https://dx.doi.org/10.17504/protocols.io.ddj24m Copy   

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Authors: Arindam Naha
Summary: For studying intracellular accumulation of CTB subunits in E coli, bacteria were grown up to 4 hours following induction by 0.2% of arabinose. Post induction, bacterial cultures were mixed with equal volume of translation-translocation halt cocktail (200 µg/ml chloramphenicol, 200 mM sodium azide, 9.5% ethanol) and immediately placed in an ice water bath for 15 minutes of incubation to arrest protein synthesis and translocation. Cells were harvested by centrifugation at 14000 rpm for 15 minutes at 4°C. Supernatants were preserved and used for CT-ELISA experiment as described before. Pellets were then suspended in TME buffer (20 mM Tris-Cl pH 8.0, 2mM β-mercaptoethanol, 1 mM EDTA), preferably 1/10th volume of the original culture and disrupted by sonication. After centrifugation at 13,000 rpm at 4°C for 15 min to remove any unbroken cells, crude cell lysates were used for measurement of protein concentration using Bradford assay (BioRad, USA). Equal amount (≥80 μg) of protein samples were mounted on to 15% SDS-PAGE. Blots were probed with classical CTB-specific monoclonal antibody (anti-Cla CTB, 1:5000) or anti-beta lactamase (anti-βla 1:500) antibody. A portion of the same blot was stained with Ponceau S staining solution (Sigma-Aldrich, St Louis, MO, USA) to validate whether equal amount protein samples were transferred in the blot. Blots were developed using West Pico Chemiluminescent Substrate (Thermo Scientific, Rockford, IL) according to the manufacturer’s instructions and visualized in ChemiDoc XRS+ system (BioRad, USA). Densitometric analysis of band intensities was performed using the Multi Gauge software V2.3(FujiFilm). .justify:after { content: ""; display:inline-block; width: 100%; }

Proper citation: Arindam Naha 2019. Western Blot CTB. protocols.io https://dx.doi.org/10.17504/protocols.io.bastieen Copy   

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