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Authors: Matthew Farnitano, Andrea Sweigart
Group: Mimulus
Summary: This is a standard PCR protocol used to amplify MgSTS markers from genomic DNA for genotyping. MgSTS markers were developed for use in Mimulus species and typically span length polymorphisms between conserved sequence regions.
Proper citation: Matthew Farnitano, Andrea Sweigart 2019. PCR protocol for MgSTS marker genotyping. protocols.io dx.doi.org/10.17504/protocols.io.baf3ibqn Copy
Authors: Joanne Kamens, Addgene The Nonprofit Plasmid Repository
Group: Re-Entering Labs Post COVID-19 Shutdown
Summary: Disclaimer: The SOP presented here has been designed by the Addgene nonprofit plasmid repository and is being shared outside of Addgene for informational purposes only. If you choose to reuse or repurpose this SOP in another location, please note that you do so at your own risk; you should ensure that any local guidance is also adhered to. None of the authors, contributors, administrators, or anyone else associated with protocols.io, can be held responsible for your use of the information contained in or linked to from these web pages.
Proper citation: Joanne Kamens, Addgene The Nonprofit Plasmid Repository 2020. Addgene Covid-19, Back To Work Training. protocols.io dx.doi.org/10.17504/protocols.io.bhukj6uw Copy
Authors: Jenn Coughlan, Jen Modliszewski, Cathy Rushworth
Group: Mimulus
Proper citation: Jenn Coughlan, Jen Modliszewski, Cathy Rushworth 2020. DNA quantification in plates (picogreen protocol). protocols.io dx.doi.org/10.17504/protocols.io.bbjdiki6 Copy
Authors: Addgene The Nonprofit Plasmid Repository
Summary: The following protocol is for bacterial transformation. To see the full abstract and additional resources, visit the Addgene protocol page.
Proper citation: Addgene The Nonprofit Plasmid Repository 2019. Bacterial transformation. protocols.io dx.doi.org/10.17504/protocols.io.4f9gtr6 Copy
Authors: Noreen Wauford
Proper citation: Noreen Wauford 2017. Cleavage Assay. protocols.io dx.doi.org/10.17504/protocols.io.hd5b286 Copy
Authors: Sherry Cryer
Summary: de novo sequencing mass spectrometry is an very important process where amino acid sequences can be interpreted from tandem mass spectra without the assistance of a database.also Peptidomics analysis studies can help yoou gain a better understanding of information about disease states, drug efficacy or toxicity.
Proper citation: Sherry Cryer 2018. de novo sequencing mass spectrometry. protocols.io dx.doi.org/10.17504/protocols.io.usxewfn Copy
Authors: Promega, Trevor Wagner
Group: Promega
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: Promega, Trevor Wagner 2018. PCR Cloning with Blue/White Selection and Easy Insert Excision. protocols.io dx.doi.org/10.17504/protocols.io.t89erz6 Copy
Authors: Qin Hong
Summary: The Handwriting Test for Preschool Children (HT-PRE), which is a newly developed handwriting screening tool for preschool children aged 5–6 years old in Mainland China, has displayed a very good internal consistency, acceptable test-retest reproducibility, and good criterion-based validity, and has also shown good application prospects for handwriting difficulty screening in a clinical setting.
Proper citation: Qin Hong 2019. Handwriting Test for Preschool Children (HT-PRE): a new tool to assess the handwriting ability of preschool children aged 5–6 years old in Mainland China. protocols.io dx.doi.org/10.17504/protocols.io.5fmg3k6 Copy
Authors: Frank Aylward
Summary: Week 1Introduction to parsing FASTA files.Commands to be entered into the command line are in bold. Here we will be using various base Unix commands such as head, tail, sort, wget, and others. We will also be using the seqkit tool to process FASTA files. The main page for seqkit is here:https://github.com/shenwei356/seqkit
Proper citation: Frank Aylward 2018. SYSB 3036 W02: Parsing FASTA files. protocols.io dx.doi.org/10.17504/protocols.io.vfwe3pe Copy
Authors: Angel Justiz-Vaillant
Summary: This reagent can be used in ELISA, Western blotting and Dot blot to detect antigens and antibodies. It is important in the immunodiagnosis of infectious diseases and other problems. Protein L binds to kappa light chains of immunoglobulins from many animal species including human, mouse, rat, chicken, hamster and pig [1].1. Justiz-Vaillant AA, Akpaka PE, McFarlane-Anderson N, Smikle MF. Comparison of techniques of detecting immunoglobulin-binding protein reactivity to immunoglobulin produced by different avian and mammalian species.West Indian Med J. 2013;62(1):12-20.
Proper citation: Angel Justiz-Vaillant 2020. Preparation of horseradish peroxidate (HRP) conjugated Peptostreptococcal protein-L by the periodate method.. protocols.io dx.doi.org/10.17504/protocols.io.bjkkkkuw Copy
Authors: Randy True
Group: FloodLAMP, XPRIZE Rapid Covid Testing
Summary: The inactivation protocol uses a chemical plus heat to break open cells (including virus, if present) and preserve the RNA. The inactivation can be done outside a lab setting, at the point of sample collection or drop-off. This is our preferred mode, and we refer to these as "inactivation stations". Prior to inactivation, the samples may contain live, contagious virus, so it is crucial that personnel use proper PPE and safe handling procedures. Many current community screening efforts are performing non-lab based sample processing (for example, Dave O'Connor and Chris Mason), and we have followed their examples with the use of splash guards in addition to PPE.After inactivation, samples should be refrigerated or stored on ice before same-day processing through the assay.On our website are our protocols in worksheet form as we use in the lab. This and more information will be live soon.
Proper citation: Randy True 2020. FloodLAMP Inactivation Protocol v3.1. protocols.io dx.doi.org/10.17504/protocols.io.bk5nky5e Copy
Authors: Katy Parker, Matt Bristow
Summary: There has been considerable interest in the measurement of cortisol in hair as a biomarker of stress since work by Davenport et al. (2006). The methodology developed by our laboratory is intended to allow for a scalable method for hair analysis that can be undertaken for large numbers of hair samples and has the minimum number of steps to reduce the chance of error and reduce costs. Our method reduces the number of steps where the sample is transferred to another container as it is kept in the same tube from initial weighing through to end of the methanol incubation. The procedure is also very scalable. The ‘rate limiting’ step for our hair analysis procedure is the grinding of the hair and this has been partially automated using the MP Biological Fast Prep grinder which is highly scalable. Final quantification of cortisol is performed via an ELISA assay, an assay which is rapid and cost effective.References:Davenport, M. D., Tiefenbacher, S., Lutz, C. K., Novak, M. a., & Meyer, J. S. (2006). Analysis of endogenous cortisol concentrations in the hair of rhesus macaques. General and Comparative Endocrinology, 147(3), 255–261. https://doi.org/10.1016/j.ygcen.2006.01.005
Proper citation: Katy Parker, Matt Bristow 2020. Hair cortisol analysis protocol. protocols.io dx.doi.org/10.17504/protocols.io.bqevmte6 Copy
Authors: E. Dale Abel
Group: Diabetic Complications Consortium
Summary: This protocol describes the procedures used by the DiaComp for evaluating mitochondrial function.Diabetic Complication:References 1. Veksler VI, Kuznetsov AV, Sharov VG, Kapelko VI, Saks VA. Mitochondrial respiratory parameters in cardiac tissue: a novel method of assessment by using saponin-skinned fibers. Biochim Biophys Acta. 1987;892:191-6. 2. Saks VA, Veksler VI, Kuznetsov AV, Kay L, Sikk P, Tiivel T, Tranqui L, Olivares J, Winkler K, Wiedemann F, Kunz WS. Permeabilized cell and skinned fiber techniques in studies of mitochondrial function in vivo. Mol Cell Biochem. 1998;184:81-100. 3. Ouhabi R, Boue-Grabot M, Mazat JP. Mitochondrial ATP synthesis in permeabilized cells: assessment of the ATP/O values in situ. Anal Biochem. 1998;263:169-75.
Proper citation: E. Dale Abel 2019. Evaluation of Mitochondrial Function. protocols.io dx.doi.org/10.17504/protocols.io.7s8hnhw Copy
Authors: Angel Justiz-Vaillant
Group: University of the West Indies, [email protected]
Summary: A recombinant protein that combines the IgG-binding domains of SpA and SpG was developed, and labelled to horseradish peroxidase. It was used as universal conjugate in ELISA for the assessment of antibodies against Brucella spp in cattle, sheep, dogs, goats and pigs. It was reported that similar results as the one shown using the chimeric protein AG were obtained when murine monoclonal antibody-enzyme conjugates were used [1,2]. References1. Justiz-Vaillant AA, Akpaka PE, McFarlane-Anderson N, Smikle MF. Comparison of techniques of detecting immunoglobulin-binding protein reactivity to immunoglobulin produced by different avian and mammalian species.West Indian Med J. 2013;62(1):12-20.2. Justiz-Vaillant AA, McFarlane-Anderson N, and Smikle M. “Bacterial Immunoglobulin (Ig)-Receptors: Past and Present Perspectives.” American Journal of Microbiological Research, vol. 5, no. 2 (2017): 44-50. doi: 10.12691/ajmr-5-2-4.
Proper citation: Angel Justiz-Vaillant 2020. Preparation of a protein-AG conjugated to horseradish peroxidase by the periodate method.. protocols.io dx.doi.org/10.17504/protocols.io.bjk6kkze Copy
Authors: Marco Cosentino, Elisa Storelli, Alessandra Luini, Massimiliano Legnaro, Emanuela Rasini, Marco Ferrari, Franca Marino
Summary: This recepe is used in the following protocols:- Separation and purification of human PBMC from FRESH BLOOD- Separation and purification of human PBMC from BUFFY COAT- Magnetic bead-based CD4+ T cell isolation from PBMCs with Dynabeads: CD4 Positive Isolation Kit- Magnetic bead-based TREG-TEFF cell isolation from PBMC with Miltenyi CD4+CD25+ Regulatory T cell Isolation Kit- Staining of human PBMC or ISOLATED SUBSETS with Cell Proliferation Dye-eFluor™ 670 (CPD-eFluor670) for cell proliferation evaluation by Flow Cytometry
Proper citation: Marco Cosentino, Elisa Storelli, Alessandra Luini, Massimiliano Legnaro, Emanuela Rasini, Marco Ferrari, Franca Marino 2020. SOLUTION- 02 - Phosphate Buffered Saline (PBS). protocols.io dx.doi.org/10.17504/protocols.io.biahkab6 Copy
Authors: Soon Kiat Lau, Jeyamkondan Subbiah
Proper citation: Soon Kiat Lau, Jeyamkondan Subbiah 2020. TDT Sandwich: An Open Source Dry Heat System for Characterizing the Thermal Resistance of Microorganisms. protocols.io dx.doi.org/10.17504/protocols.io.36agrae Copy
Authors: Chang-Ming Bai, Lu-Sheng Xin, Umberto Rosani, Biao Wu, Qing-Chen Wang, Xiao-Ke Duan, Zhi-Hong Liu, Chong-Ming Wang
Group: GigaScience Press
Summary: Background: The blood clam, Scapharca (Anadara) broughtonii, is an economically and ecologically important marine bivalve of the Family Arcidae. The efforts that have been made to study their population genetics, breeding, cultivation and stock enrichment were somewhat hindered by the lack of a reference genome. Here, we reported the complete genome sequence of S. broughtonii, a first reference genome of the Family Arcidae.Funding: A total of 75.79 Gb clean data was generated with the PacBio and Oxford Nanopore platforms, which represented approx. 86× coverage of the S. broughtonii genome. De novo assembly of these long reads resulted in an 884.5 Mb genome, with a contig N50 of 1.80 Mb and scaffold N50 of 45.00 Mb, respectively. Genome Hi-C scaffolding resulted in 19 chromosomes containing 99.35% of bases of the assembled genome. Genome annotation revealed that a considerable part of the genome (46.1%) is composed by repeated sequences, while 24,045 protein-coding genes were predicted and 84.7% of them were annotated.Conclusion: We report here the chromosomal-level assembly of the S. broughtonii genome based on long read sequencing and Hi-C scaffolding. The genomic data could be served as reference genome for the Arcidae Family and will provide a valuable resource for the scientific community and aquaculture sector.
Proper citation: Chang-Ming Bai, Lu-Sheng Xin, Umberto Rosani, Biao Wu, Qing-Chen Wang, Xiao-Ke Duan, Zhi-Hong Liu, Chong-Ming Wang 2019. Key protocols for chromosome-level genome assembly of the Scapharca (Anadara) broughtonii.. protocols.io dx.doi.org/10.17504/protocols.io.zimf4c6 Copy
Authors: Julie Jerber, James Haldane, Juliette Steer, Daniel Pearce, Minal Patel
Group: Cellular Generation and Phenotyping
Summary: This protocol describes a method for the production of dopaminergic neurons from human iPSCs, using a 52-day long differentiation method adapted from doi.org/10.1038/nature10648. Steps in this protocol assume the use of a single 12 well plate for the purposes of differentiation; adjust volumes accordingly depending on labware used.
Proper citation: Julie Jerber, James Haldane, Juliette Steer, Daniel Pearce, Minal Patel 2020. Generation of iPSC-derived dopaminergic neurons. protocols.io dx.doi.org/10.17504/protocols.io.bjpgkmjw Copy
Authors: Brandt Pence
Proper citation: Brandt Pence 2017. Isolation of Monocytes from Whole Blood With Magnetic Negative Sorting. protocols.io dx.doi.org/10.17504/protocols.io.kwtcxen Copy
Authors: Kristin Corrier/Nathan VerBerkmoes
Group: VERVE Net, Sullivan Lab
Summary: For use in the "FASP Kit Protocol-ORNL Developed for Bacteriophage"
Proper citation: Kristin Corrier/Nathan VerBerkmoes 2016. Digestion Solution. protocols.io dx.doi.org/10.17504/protocols.io.dek3cv Copy
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