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Authors: Leonardo Caserta
Group: Coronavirus Method Development Community, Diel Lab

Proper citation: Leonardo Caserta 2021. First-strand synthesis and touchdown PCR for SARS-CoV-2. protocols.io https://dx.doi.org/10.17504/protocols.io.br54m88w Copy   

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Authors: Aldo Solari, Sylvia Ortiz, Camila Ihle-Soto, Eduardo Costoya, Antonella Bacigalupo, Juana Paola Correa
Summary: DNA-Blot genotyping of Trypanosoma cruzi Discrete Typing Units (DTU) with radiolabeled  probes

Proper citation: Aldo Solari, Sylvia Ortiz, Camila Ihle-Soto, Eduardo Costoya, Antonella Bacigalupo, Juana Paola Correa 2018. DNA-Blot genotyping of Trypanosoma cruzi Discrete Typing Units (DTU) with radiolabeled probes. protocols.io https://dx.doi.org/10.17504/protocols.io.sz2ef8e Copy   

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Authors: Yingchao Xue, Xiping Zhan, Shisheng Sun, Senthilkumar S. Karuppagounder, Shuli Xia, Valina L Dawson, Ted M Dawson, John Laterra, Jianmin Zhang, Mingyao Ying
Group: Neurodegeneration Method Development Community

Proper citation: Yingchao Xue, Xiping Zhan, Shisheng Sun, Senthilkumar S. Karuppagounder, Shuli Xia, Valina L Dawson, Ted M Dawson, John Laterra, Jianmin Zhang, Mingyao Ying 2020. Protocols for Synthetic mRNAs Drive Highly Efficient iPS Cell Differentiation to Dopaminergic Neurons. protocols.io https:// Copy   

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Authors: Jingrong Ye
Summary: We analyzed the demographic, clinical, and virological data of residents newly diagnosed with HIV in Beijing. We did population-based sequencing of the pol gene on plasma specimens and identified drug resistance mutations using the World Health Organization (WHO) list for surveillance of drug resistance mutations. HIV-1 subtype analyses utilized the automated subtyping tool COMET.

Proper citation: Jingrong Ye 2019. Protocol of HIV TDR and Subtype test in Beijing. protocols.io https://dx.doi.org/10.17504/protocols.io.8ahhsb6 Copy   

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Authors: João Vitor Molino
Summary: This protocols describe the steps to perform a western blot in Chlamydomonas reinhardtii cell lysate and supernatant samples.

Proper citation: João Vitor Molino 2018. Western blotting in Chlamydomonas reinhardtii. protocols.io https://dx.doi.org/10.17504/protocols.io.kfpctmn Copy   

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Authors: Sarah Giuliani
Group: Coleman Lab
Summary: Experiment purpose is to monitor the time-course of a large-scale infection of host cyanobacteria by phage under variable media conditions and obtain samples for proteomic and transcriptomic analysis.9 Hourly Timepoints: 0, 1, 2, 3, 4, 6, 8, 10, 14Sampling is to collect supernatant fractions to determine total dissolved phosphate and soluble reactive phosphate (TDP/SRP) for analysis of inorganic phosphate concentration and phosphate in dissolved organic material (DOM); and to collect pellets to isolate RNA for transcriptomics and proteins for proteomics.

Proper citation: Sarah Giuliani 2016. Sampling for RNA / Protein/ DOM/ PO4: Large-scale One-step Phage Infection of Cyanobacteria. protocols.io https://dx.doi.org/10.17504/protocols.io.fkvbkw6 Copy   

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Authors: Bonnie Poulos, Jenn Brum, Elke Allers, Christine Schirmer
Group: VERVE Net, Sullivan Lab
Summary: This protocol gives a method for treatment and preservation of seawater samples that will be subject to fluorescence in situ hybridization (FISH). Samples are used with fluorescent probes to determine what populations of bacteria or other microbes are present. The filters may also be used to simply count the number of microbes in a sample. Requires varying amounts of 10µm or 200µm filtrate (up to 11.5L for open ocean depth profile) as shown in Table I.

Proper citation: Bonnie Poulos, Jenn Brum, Elke Allers, Christine Schirmer 2015. Treatments and Preservation of Seawater Samples for FISH. protocols.io https://dx.doi.org/10.17504/protocols.io.c5by2m Copy   

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Authors: Mark Dewitt, Julia Wong
Group: Innovative Genomics Institute, CornLab
Summary: sgRNA template assembly, in vitro T7 transcription, and SPRI bead cleanup

Proper citation: Mark Dewitt, Julia Wong 2016. In vitro transcription of guide RNAs. protocols.io https://dx.doi.org/10.17504/protocols.io.exabfie Copy   

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Authors: Tim Mercer
Summary: RNA sequencing can measure both gene or isoform expression, and reconstruct novel and complex spliced isoforms. However, the sheer size and complexity of the transcriptome, as well as technical bias, can confound analysis with RNA-seq. To assess the impact of these variables, we have developed a set of RNA sequins that represent synthetic genes that act as internal controls during RNA sequencing.Each RNA sequin represents an individual isoform, with multiple isoforms forming artificial gene loci that are encoded within the in silico chromosome (chrIS). By modulating the relative abundance of individual sequin isoforms we can emulate alternative splicing, whilst modulating the abundance of multiple isoforms we can emulate gene expression. Accordingly, RNA sequins are mixed at different concentration to emulate differences in gene expression and alternative splicing. By sequentially diluting sequins, we can establish a reference ladder across a range of gene expressions. We formulate multiple alternative mixtures that differ in the concentration of individual sequins. By comparing mixtures, we can emulate differential gene expression and alternative splicing between samples. By contrast, RNA sequins with invariant concentrations between mixtures provide static scaling factors that enable quantitative normalization between multiple RNAseq libraries.The RNA sequin mixture is added to a user’s RNA sample at a fractional concentration prior to library preparation. The combined sample and the sequins then together undergo sequencing. The sequins can then be distinguished in the output library by their synthetic sequence, and analyzed as internal controls.For further detailed background on the design, validation and use of sequins, we refer users to ‘Spliced synthetic genes as internal controls in RNA sequencing experiments’ by Hardwick et al., (2016) NAture Methods DOI:10.1038/nmeth.3958\

Proper citation: Tim Mercer 2019. Addition of RNA sequins to sample for RNA sequencing.. protocols.io https://dx.doi.org/10.17504/protocols.io.x8cfrsw Copy   

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Authors: Eliseos Mucaki, Peter Rogan
Summary: Accurate radiation dose estimates are critical for determining eligibility for therapies by timely triaging of exposed individuals after large-scale radiation events. However, the universal assessment of a large population subjected to a nuclear spill incident or detonation is not feasible. Even with high-throughput dosimetry analysis, test volumes far exceed the capacities of first responders to measure radiation exposures directly, or to acquire and process samples for follow-on biodosimetry testing. We aimed to design a protocol which can significantly reduce data acquisition and processing requirements for timely treatment of eligible, affected individuals in population-scale radiation exposures.Method in Brief: The spatial boundaries of graduated radiation exposures were determined by targeted, multistep geostatistical analysis of small population samples. Physical radiation plumes modelled nuclear detonation scenarios of simulated exposures at 22 US locations. Models assumed only location of the epicenter and historical, prevailing wind directions/speeds. Initially, locations proximate to these sites were randomly sampled (0.1% of population). Empirical Bayesian kriging established radiation dose contour levels circumscribing these sites. Densification of each plume identified critical locations for additional sampling. After repeated kriging and densification, overlapping grids between each pair of contours of successive plumes were compared based on their diagonal Bray-Curtis distances and root-mean-square deviations, which provided criteria (

Proper citation: Eliseos Mucaki, Peter Rogan 2020. Protocol for Geostatistical Determination of Radiation Dosimetry Maps of Population-Scale Exposures. protocols.io https://dx.doi.org/10.17504/protocols.io.ba4nigve Copy   

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Authors: Kholoud Refaat Ahmed, Amal Hassan Abdel Rahman, Manar Abdul Waniss Mohammed Abdul Aziz

Proper citation: Kholoud Refaat Ahmed, Amal Hassan Abdel Rahman, Manar Abdul Waniss Mohammed Abdul Aziz 2018. Molecular Assessment of Kallikrein 3 in Ameloblastoma and Odontogenic Keratocyst (In-vitro Study). protocols.io https://dx.doi.org/10.17504/protocols.io.uazesf6 Copy   

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Authors: Pip Wilson, Diep Ganguly, Xin Hou, Barry Pogson
Group: Pogson Genomics Group
Summary: Extraction of genomic DNA from Arabidopsis leaf material.

Proper citation: Pip Wilson, Diep Ganguly, Xin Hou, Barry Pogson 2018. CTAB genomic DNA extraction from Arabidopsis leaf material. protocols.io https://dx.doi.org/10.17504/protocols.io.quidwue Copy   

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Authors: Jiaxin Li

Proper citation: Jiaxin Li 2020. Single digested. protocols.io https://dx.doi.org/10.17504/protocols.io.bni5mcg6 Copy   

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Authors: Joachim Nwezeobi, Onyeyirichi Onyegbule, Chukwuemeka Nkere, Joseph Onyeka, Sharon van Brunschot, Susan Seal, John Colvin

Proper citation: Joachim Nwezeobi, Onyeyirichi Onyegbule, Chukwuemeka Nkere, Joseph Onyeka, Sharon van Brunschot, Susan Seal, John Colvin 2020. Whitefly DNA extraction, partial mtCO1 gene amplification and gel electrophoresis. protocols.io https://dx.doi.org/10.17504/protocols.io.bdvri656 Copy   

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Authors: Celeste Karch, Rita Martinez, Jacob Marsh
Group: Neurodegeneration Method Development Community

Proper citation: Celeste Karch, Rita Martinez, Jacob Marsh 2019. Serial Dilution of Nucleofected iPSC Pools. protocols.io https://dx.doi.org/10.17504/protocols.io.x76frre Copy   

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Authors: Niklas Hausmann, Amy Prendergast, Andreas Lemonis, Jana Zech, Patrick Roberts, Panagiotis Siozos, Demetrios Anglos
Summary: Elemental analysis of biogeochemical archives is an established technique used to study climate in a range of applications, including ocean circulation, glacial/interglacial climates, and anthropogenic climate change. Data from mollusc archives are especially important because of their global abundance and sub-annual resolution. Despite this potential, they are underrepresented among palaeoclimate studies, due to enigmatic physiological influences skewing the elemental record. Understanding the patterns behind these influences will improve data interpretation and lead to the development of new climate proxies. Here, we show for the first time that extensive spatial mapping of multiple mollusc specimens using Laser Induced Breakdown Spectroscopy (LIBS) across a wider region can resolve enigmatic patterns within the elemental record caused by physiological influences. 2D elemental (Mg/Ca) maps of whole limpet shells (Patella caerulea) from across the Mediterranean revealed patterns of variability within individual mollusc records as well as within isochronous parts of specimens. By registering and quantifying these patterns, we established previously uninterpretable correlations with temperature (R2 > 0.8, p

Proper citation: Niklas Hausmann, Amy Prendergast, Andreas Lemonis, Jana Zech, Patrick Roberts, Panagiotis Siozos, Demetrios Anglos 2019. LIBS Mapping of Mg/Ca ratios in marine mollusc shells. protocols.io https://dx.doi.org/10.17504/protocols.io.zc9f2z6 Copy   

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Authors: Alan Cone
Group: Ju Lab
Summary: Cleans and concentrates any DNA. Typically useful after PCR, restriction digest, or anything where DNA was altered and now you need your newly altered DNA in a pure form.

Proper citation: Alan Cone 2016. Zymo Research DNA Clean & Concentrator. protocols.io https://dx.doi.org/10.17504/protocols.io.egbbbsn Copy   

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Authors: Jason Kim
Group: Mouse Metabolic Phenotyping Centers
Summary: This experiment involves a spectrophotometric measurement using Roche Cobas Clinical Chemistry Analyzer. Serum levels of creatinine reflect systemic protein metabolism.

Proper citation: Jason Kim 2019. U Mass - Creatinine. protocols.io https://dx.doi.org/10.17504/protocols.io.xvjfn4n Copy   

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Authors: Jay Hesselberth, Amanda Richer
Summary: Methods to measure heterogeneity among cells are rapidly transforming our understanding of biology but are currently limited to static measurements of DNA, RNA, and protein abundance. We developed an approach to simultaneously measure biochemical activities and mRNA abundance in single cells to understand the heterogeneity of DNA repair across thousands of human lymphocytes, identifying known and novel cell-type-specific DNA repair phenotypes. Our method provides a general framework for understanding functional heterogeneity among single cells.

Proper citation: Jay Hesselberth, Amanda Richer 2019. Single-cell analysis of functional heterogeneity in DNA repair capacity. protocols.io https://dx.doi.org/10.17504/protocols.io.uhyet7w Copy   

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Authors: G-Biosciences
Group: G-Biosciences
Summary: This protocol is part of the FOCUS™ Mitochondria Kit collection. Please refer to the appropriate protocol depending on your application.

Proper citation: G-Biosciences 2016. Isolation of Mitochondria from Soft Tissues (Liver or Brain) using the FOCUS™ Mitochondria Kit. protocols.io https://dx.doi.org/10.17504/protocols.io.e89bhz6 Copy   

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