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Authors: luanaborba
Group: L. P. Borba-Santos
Proper citation: luanaborba 2020. Determination of minimum inhibitory concentration values (MICs) against Sporothrix brasiliensis and Sporothrix schenckii. protocols.io dx.doi.org/10.17504/protocols.io.bjfbkjin Copy
Authors: Kristoffer Bach Falkenberg, Maja Rennig, Cristina Hernandez Rollan, Andreas Birk Bertelsen, Morten Norholm
Proper citation: Kristoffer Bach Falkenberg, Maja Rennig, Cristina Hernandez Rollan, Andreas Birk Bertelsen, Morten Norholm 2020. LyGo protocol collection. protocols.io dx.doi.org/10.17504/protocols.io.9nvh5e6 Copy
Authors: Bernardo Mainou
Summary: Indirect immunofluorescence infectivity assay for reovirus using LICOR Imaging System
Proper citation: Bernardo Mainou 2017. Fluorescent Focus Unit Assay using LICOR Imaging System. protocols.io dx.doi.org/10.17504/protocols.io.kqicvue Copy
Authors: Michael S. Fernandopulle, Ryan Prestil, Christopher Grunseich, Chao Wang, Li Gan, Michael E. Ward
Group: Neurodegeneration Method Development Community
Summary: Accurate modeling of human neuronal cell biology has been a long-standing challenge. However, methods to differentiate human induced pluripotent stem cells (iPSCs) to neurons have recently provided experimentally tractable cell models. Numerous methods that use small molecules to direct iPSCs into neuronal lineages have arisen in recent years. Unfortunately, these methods entail numerous challenges, including poor efficiency, variable cell type heterogeneity, and lengthy, expensive differentiation procedures. We recently developed a new method to generate stable transgenic lines of human iPSCs with doxycycline-inducible transcription factors at safe-harbor loci. Using a simple two-step protocol, these lines can be inducibly differentiated into either cortical (i3Neurons) or lower motor neurons (i3LMN) in a rapid, efficient, and scalable manner (Wang et al., 2017). In this manuscript, we describe a set of protocols to assist investigators in the culture and genetic engineering of iPSC lines to enable transcription factor-mediated differentiation of iPSCs into i3Neurons or i3LMNs, and we present neuronal culture conditions for various experimental applications. © 2018 by John Wiley & Sons, Inc.
Proper citation: Michael S. Fernandopulle, Ryan Prestil, Christopher Grunseich, Chao Wang, Li Gan, Michael E. Ward 2019. Transcription Factor-Mediated Differentiation of Human iPSCs into Neurons. protocols.io dx.doi.org/10.17504/protocols.io.54rg8v6 Copy
Authors: Michael Hagemann-Jensen, Christoph Ziegenhain, Ping Chen, Daniel Ramsköld, Gert-Jan Hendriks, Anton J.M Larsson, Omid R. Faridani, Rickard Sandberg
Proper citation: Michael Hagemann-Jensen, Christoph Ziegenhain, Ping Chen, Daniel Ramsköld, Gert-Jan Hendriks, Anton J.M Larsson, Omid R. Faridani, Rickard Sandberg 2020. Smart-seq3 Protocol. protocols.io dx.doi.org/10.17504/protocols.io.bbpyimpw Copy
Authors: Integrated Islet Distribution Program
Group: Integrated Islet Distribution Program, Integrated Islet Distribution Program and Human Islet Phenotyping Program
Summary: This SOP defines a standardized method for packaging and cold shipping of research quality islets to approved investigators of human isolated islet preparations, for use in the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) sponsored research in the Integrated Islet Distribution Program (IIDP). This protocol is written to assist the participating islet isolation centers and investigators who are part of this program.Note: This SOP was developed based on the Prodo Labs, Inc. shipping protocol and results from preliminary studies conducted by the IIDP and commissioned by the original IIDP Project Officer, and External Evaluation Committee (EEC).It was commissioned due to problems with acquiring the supplies that were used in IIDP SOP: SHP-001 and with the hope that this method is a better means for transportation of IIDP islets. Preliminary studies proved the islets by the Prodo Labs’ method were statistically as good as the original IIDP method, it was preferred by the test researchers, and was much more cost effective for the IIDP. This new method may be modified as future methods are tested and approved by the IIDP Team, Project Scientist (PS), the Program Official (PO), and EEC.
Proper citation: Integrated Islet Distribution Program 2020. Islet Culture and Preparation for Cold Shipping. protocols.io dx.doi.org/10.17504/protocols.io.bfsfjnbn Copy
Authors: Ian Mackay, Mitchell Finger, Michael Lyon, Judy Northill
Group: Public Health Virology, Forensic and Scientific Services
Summary: This protocol was developed at this laboratory but had not been previously published.
The protocol aims explicitly to amplify hepatitis A virus (HAV) strains and not other virus species. The assay targets the 5' untranslated region and is designed as a qualitative test for investigating suspected human cases of HAV infection.
Proper citation: Ian Mackay, Mitchell Finger, Michael Lyon, Judy Northill 2018. Hepatitis A MGB TaqMan. protocols.io dx.doi.org/10.17504/protocols.io.rk3d4yn Copy
Authors: Kenneth Schackart, Kattika Kaarj
Group: 481b Laboratory
Summary: This protocol details how to stain mammalian cells cultured on a 96-well plate. Actin filaments, focal adhesion sites (as indicated by the presence vinculin), and nuclei will be stained.
Proper citation: Kenneth Schackart, Kattika Kaarj 2019. Mammalian Cell Staining. protocols.io dx.doi.org/10.17504/protocols.io.xs7fnhn Copy
Authors: Sam Li
Group: BioLegend
Summary: BioLegend MojoSort™ nanobeads work in commonly used separation columns, based on our internal research as well as validation by external testing by academic labs. This simple protocol consists of following the MojoSort™ protocol to label the cells with pre-diluted MojoSort™ reagents and using the columns as indicated by the manufacturer.Note: Due to the properties of our beads, it may be possible to use far fewer beads and less antibody cocktail that with other commercial suppliers. We recommend a titration to find the best dilution factor. However, as a general rule, dilutions ranging from 1:2 to 1:10 for the antibody cocktail can be used. Dilutions ranging from 1:5 to 1:20 for the Streptavidin Nanobeads can be used. Please contact BioLegend Technical Service ([email protected]) if further assistance is needed.
Proper citation: Sam Li 2019. MojoSort™ Human NK Cell Isolation Kit Protocol. protocols.io dx.doi.org/10.17504/protocols.io.7bihike Copy
Authors: Dennis Dienst
Group: CyanoWorld
Proper citation: Dennis Dienst 2017. RNA extraction from Synechocystis sp. PCC 6803 with Trizol reagent. protocols.io dx.doi.org/10.17504/protocols.io.j3scqne Copy
Authors: Ibrahim El-khalil Adam
Summary: The author describes a new image analysis algorithm that enables identification of how many DNA fragments co-migrate during PFGE. The method is named factor of co-migration based on exponential correlation between single-fragment bands and their pixel densities "FCM-ECSB".
Proper citation: Ibrahim El-khalil Adam 2021. Calculating number of Co-Migrated DNA Fragments across Pulsed Field gel electrophoresis (PFGE) profiles: Image analysis algorithm. protocols.io dx.doi.org/10.17504/protocols.io.bthtnj6n Copy
Authors: Dr. Steven Wilhelm, Samantha Coy
Group: The Aquatic Microbial Ecology Research Group - AMERG (The Buchan, Zinser and Wilhelm labs), Samantha R Coy's Protocols
Summary: Please contact Dr. Steven Wilhelm ([email protected]) or Samantha Coy ([email protected]) for additional information regarding this protocol.
De-MetaST-BLAST, written in C++, is a script that can validate degenerate primers by performing in silico PCR amplification of the primers against metagenomic and metatranscriptomic databases (e.g. CAMERA; MG-RAST). The output is a list of in-silco PCR amplicons that are then blasted against GenBank to determine the most homologous sequence to the amplicon.
Modified from Gulvik, C. A., Effler, T. C., Wilhelm, S. W., & Buchan, A. (2012). De-MetaST-BLAST: A Tool for the Validation of Degenerate Primer Sets and Data Mining of Publicly Available Metagenomes. PLoS ONE,7(11). doi:10.1371/journal.pone.0050362
Proper citation: Dr. Steven Wilhelm, Samantha Coy 2017. Using De-MetaST-BLAST. protocols.io dx.doi.org/10.17504/protocols.io.ii2ccge Copy
Authors: Gregory Harhay
Summary: The virulence and pathogenicity of bacterial pathogens are related to their adaptability to changing environments. One process enabling adaptation is based on minor changes in genome sequence, as small as a few base pairs, within segments of genome called simple sequence repeats (SSRs) that consist of multiple copies of a short sequence (from one to several nucleotides), repeated in series. SSRs are found in eukaryotes as well as prokaryotes, and variation in them occurs at frequencies up to a million-fold higher than the average bacterial mutation rate through a process of slipped stranded mispairing (SSM) by DNA polymerase during replication. The characterization of SSR length by standard sequencing methods is complicated by the appearance of length variation introduced during the sequencing process that does not accurately quantify lower-abundance repeat number variants in a population. Here we report a computational approach to correct for process-induced artifacts, validated for tetranucleotide repeats by use of synthetic constructs of fixed, known length. We apply this method to a laboratory culture ofHistophilus somni, prepared from a single colony, and demonstrate that the culture consists of populations of distinct sequence phase and read length variants at individual tetranucleotide SSR loci.Input requirements: Closed Genome - It is recommended that only organisms with closed genomes be the subject of the analyses described here. Mapping repetitive reads to to contigs of non-closed genomes may map to multiple locations, complicating tha analysis. Mapping CCS (circular consensus sequence) wiith repetitve sequence to closed genomes are guaranted to map to a single locus if sufficent unique flanking sequence is used to confim the unique mapping. Consequently, long CCS with high base quality are the most desirable input into this workflow.
Proper citation: Gregory Harhay 2019. Steps to Create FASTQ of CCS Overlapping Genomic SSR - CCS ROI . protocols.io dx.doi.org/10.17504/protocols.io.9i7h4hn Copy
Authors: Ken Christensen, Promega, Trevor Wagner
Summary: Protocol for PCR Cloning with Blue/White Selection and Easy Insert Excision using pGEM®-T Easy Vector Systems.(For use with A3600, A3610, A1360, or A1380.)
Proper citation: Ken Christensen, Promega, Trevor Wagner 2020. PCR Cloning with Blue/White Selection--CHEM 584. protocols.io dx.doi.org/10.17504/protocols.io.bmtzk6p6 Copy
Authors: Aharon Levy, Michael Krakovsky
Summary: The objective of this protocol is to present the details of the procedures for evaluating test items’ (such as small molecules, biologics, and others) efficacy in promoting stroke rehabilitation, in the rat transient Middle Cerebral Carotid Occlusion (t-MCAO) model.
Proper citation: Aharon Levy, Michael Krakovsky 2020. EVALUATION OF TEST ITEMS EFFICACY IN PROMOTING STROKE REHABILITATION IN THE RAT STROKE MODEL (t-MCAO).. protocols.io dx.doi.org/10.17504/protocols.io.bf69jrh6 Copy
Authors: Yoan Coudert
Group: CoudertLab
Proper citation: Yoan Coudert 2020. DH5α bacteria transformation protocol. protocols.io dx.doi.org/10.17504/protocols.io.bghijt4e Copy
Authors: Anastasios Tsaousis, Eleana Kazana, Tobias von der Haar
Summary: Maps and nucleotide sequences of the Naegleria gruberi transfection plasmids (pNaeg-HYG and pNaeg-NEO) reported in Faktorová D. et al.
Proper citation: Anastasios Tsaousis, Eleana Kazana, Tobias von der Haar 2019. Naegleria gruberi plasmids. protocols.io dx.doi.org/10.17504/protocols.io.7w4hpgw Copy
Authors: BioLegend, Inc.
Group: BioLegend
Proper citation: BioLegend, Inc. 2016. Western Blotting Protocol. protocols.io dx.doi.org/10.17504/protocols.io.e2abgae Copy
Authors: andrei.herdean
Group: Climate Change Cluster
Proper citation: andrei.herdean 2021. Action Spectra protocol for Opentrons OT-1 liquid handling robot . protocols.io dx.doi.org/10.17504/protocols.io.br6vm9e6 Copy
Authors: Paloma Varela
Proper citation: Paloma Varela 2019. Fluorescence-based Thermal Shift Assay (TSA). protocols.io dx.doi.org/10.17504/protocols.io.vg2e3ye Copy
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