eamag/protocols-with-steps huggingface dataset:
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
This protocol describes the data collection and analysis performed in "Invasion genetics of the silver carp (Hypophthalmichthys molitrix) across North America: Differentiation of fronts, introgression, and eDNA detection". Published in PLOS One, 2018.

The invasive silver carp Hypophthalmichthys molitrix escaped from southern U.S. aquaculture during the 1970s to spread throughout the Mississippi River basin and steadily moved northward, now reaching the threshold of the Laurentian Great Lakes. The silver carp is native to eastern Asia and is a large, prolific filter-feeder that decreases food availability for fisheries. The present study evaluates its population genetic variability and differentiation across the introduced range using 10 nuclear DNA microsatellite loci, sequences of two mitochondrial genes (cytochrome b and cytochrome c oxidase subunit 1), and a nuclear gene (ribosomal protein S7 gene intron 1). Populations are analyzed from two invasion fronts threatening the Great Lakes (the Illinois River outside Lake Michigan and the Wabash River, leading into the Maumee River and western Lake Erie), established areas in the southern and central Mississippi River, and a later Missouri River colonization. Results discern considerable genetic diversity and some significant population differentiation, with greater mtDNA haplotype diversity and unique microsatellite alleles characterizing the southern populations. Invasion fronts significantly differ, diverging from the southern Mississippi River population. About 3% of individuals contain a unique and very divergent mtDNA haplotype (primarily the southerly populations and the Wabash River), which may stem from historic introgression in Asia with female largescale silver carp H. harmandi. Nuclear microsatellites and S7 sequences of the introgressed individuals do not significantly differ from silver carp. MtDNA variation is used in a high-throughput sequence assay that identifies and distinguishes invasive carp species and their population haplotypes (including H. molitrix and H. harmandi) at all life stages, in application to environmental (e)DNA water and plankton samples. We discerned silver and bighead carp eDNA from 10 bait shops in the Great Lakes watershed, indicating that release from retailers comprises another likely vector. Our findings provide key baseline population genetic data for understanding and tracing the invasion’s progression, facilitating detection, and evaluating future trajectory and adaptive success.


[STEPS]
SECTION: Tissue sample collection & DNA extraction
?. 
SECTION: DNA Sequencing
?. 
SECTION: Microsatellite Data Collection
?. 
SECTION: eDNA sample collection
?. 
SECTION: eDNA library preparation
?. 
SECTION: custom eDNA analysis scripts
?.
TITLE: Snap freezing brains
AUTHORS: Molly Brennan, Olivier George

[STEPS]
?. Snap freezing brains euthanize animal in either a bell jar of Isoflorane or in C02 chamber.once animal is dead you then can use scissors and or a gullitoine to remove head at nape of neck
?. Brain Removal-Use a pair of scissors to cut connective tissue under the skin, reaching in between the eyes-Cut skin open, exposing muscles on top of skull to the eyes-Use scalpel, cut muscle connecting to the skull bone on both sides-Carefully place bone cutter against the inside of the skull, and make cuts following the scalpel incision-Lift the occipital, parietal, and frontal bones using forceps, fully exposing the brain-Cut membrane on top and scoop brain out of the skull; cut optical nerves and connective tissue
?. Once brain is removed place it in -38-40 degree tempeture of Ispentane and or 2 methylbutane
?. Once brain is submerged into the 2 methylbutane it needs to stay there for 30-45 seconds or until brain is frozendo not have temp lower than -40 degrees or the brain will crack and or split in two. VERY IMPORTANT
?. take brain out and place in foil with a label or vial. your choice. store in -80 freezer.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
Hypertension is a long-term medical condition in which the pressure of blood travelling through the arteries is too high. This puts increased strain on the heart which, if left untreated, can lead to increased risk of heart attack or stroke. It is extremely common and is a major cause of death and disease worldwide, however many cases remain undiagnosed and treatment is not as effective as it could be. General practitioners play an important role in the early diagnosis and treatment of high blood pressure. In general, patients are treated with a combination of different drugs, as using a single drug is only effective in a limited number of patients. If initial treatment with two drugs fails then the current guidelines recommend increasing the dosage of these drugs, switching to two different drugs or adding a third drug. If treatment with three drugs is needed, the guidelines favour the use of fixed-dose or single-pill combinations, because reducing multiple drug treatment to one tablet daily helps improve the chance of patients always remembering to take their medication. The aim of this study is to complete a large-scale survey to investigate how well blood pressure is controlled in hypertensive patients who are being treated with more than one blood pressure medication.

[STEPS]
?.
TITLE: Diagnostic Restriction Digest (Instructor Protocol)
AUTHORS: Brian    Teague

[DESCRIPTION]
This is the instructor protocol for
 


[STEPS]
SECTION: Setup
1. Aliquot the  : 20 µL ul aliquots, 1 per 4 students

SECTION: Setup
2. Aliquot the   enzyme: 4 µL of enzyme in 16 µL of  , 1 per 4 students.

SECTION: Instructor Tips &amp; Common Student Errors
3. Instructor Tips
In a more advanced course, I would let students select which enzyme to use (chosen from the ones in the freezer.) In an intro course, I choose the enzyme for them. PvuII is a good choice because it has two cut sites in the backbone and one in the GFP insert. This makes it easy to distinguish between the correct plasmid and one with a GFP still present (maybe not glowing because of a mutation? Or some other error?)

PvuII is a little on the expensive side -- but if you're giving the digest a full hour, diluting down to 2 units per ul gives two advantages: it decreases the amount used, and increases the volume of the pipetting step. Make sure you use the correct diluent, though.

I used to include a heat-inactivation step. I don't any more because the SDS in the purple loading dye denatures the enzyme.

Sometimes a student's miniprep isn't concentrated enough to get a full microgram of DNA into the digest. As long as they can get at least 500 ng in the digest, that should be enough to see on a gel. It's more acceptable to decrease the mass of DNA than it is to increase the volume -- contaminants in the miniprep often get in the way of the digest, particularly leftover ethanol.

I don't have a positive control in this experiment as written. Maybe add one?
SECTION: Instructor Tips &amp; Common Student Errors
4. Common Student Errors
Not mixing the reaction well.
Not loading the entire reaction onto the gel. (Because the first one they did was a PCR, it's easy to assume that you just need 1-2 ul without reading the protocol carefully.)
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>This protocol is used for performing Type IIS assembly by either <em>BsaI</em> or <em>SapI</em>-mediated restriction/ligation using Loop assembly.</p>
<p> </p>
<p>Loop assembly comprises 8 receiver plasmids in odd and even levels (4 per level), which contain directional overhangs for higher level assembly. Odd and even level receiver plasmids are used in a recursive schema for assembly in iterative 'loops'. Any assembly (except for L0 part composition into a L1 transcriptional unit) requires the usage of 4 donor plasmids (each in a specific position) and one receiver plasmid. <em>BsaI</em> is used for assembly into odd levels using kanamycin selection, and <em>SapI</em> is used in even levels using spectynomycin selection. Loop odd receivers use the Common Syntax standard for L0 part assembly into TUs (Patron, <em>et al.</em> 2015 [10.1111/nph.13532]). Then, 4 L1 plasmids (positions 1-4) are assembled into an even receiver plasmid to provide a L2 assembly (4 TUs). For higher level assemblies the same assembly structure is followed, 4 L2 plasmids (positions 1-4) with an odd receiver to yield a L3 assembly (16 TUs).</p>
<p> </p>
<p>The Loop type IIS assembly protocol is based on: </p>
<p class="p1"><span class="s1">“Patron, NJ (2016) DNA Assembly for Plant Biology. Current Protocols in Plant Biology 1:1-13 [doi: 10.1002/cppb.20038]”, but with slight modifications for DNA concentrations:</span></p>
<p> </p>
<p><strong>This protocol uses a target amount of each donor DNA of 15 fmol and 7.5 fmol for receiver plasmid DNA in a 10 µL reaction. In order to perform accurate dispensing, the plasmid DNA is diluted to their corresponding concentration of 15 fmol/µL for donor parts and of 7.5 fmol/µL for the receiver plasmid, and then 1 µL of each plasmid is added to the DNA mix. The plasmid part target concentration (in ng/uL) equals to each donor plasmid length / 100, and of the receiver plasmid length / 200. </strong></p>
<p> </p>

[BEFORE_START]
<p>Thaw 10X T4 ligase buffer at RT and then leave on ice. Keep BSA on ice. Briefly vortex 10X ligase buffer before using.</p>

[GUIDELINES]
<p>Each level (odd and even) contains four 'receiver' plasmids:</p>
<p> </p>
<p>pOdd-1, 2, 3, 4</p>
<p>pEven-1, 2, 3, 4</p>
<p> </p>
<p>For a the assembly of L0 parts into a L1 TU, a variable number of parts can be used for the asssembly of the TU, however, for higher level assemblies only 4 donor modules can be assembled into any receiver.</p>
<p> </p>
<p>Use only high-purity plasmid preparations. Genomic contamination of plasmid DNA will interfere with assembly reactions.</p>

[STEPS]
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?.
TITLE: DEMoNS protocol for measurement and analysis of eye movements
AUTHORS: Jenny  Nij Bijvank

[DESCRIPTION]
For questions, send an email to info.demonsprotocol@gmail.com or vanrijn@amsterdamumc.nl
The complete protocol and instructions on measurement and analysis of eye movements with the DEMoNS protocol.

Quantitative saccadic testing is a non-invasive method of evaluating the neural networks involved in the control of eye movements. The DeMONS protocol is a standardized and reproducible protocol for infrared oculography measurements of eye movements and analysis, which can be applied for various diseases in a multicenter setting. In the accompanying manuscript in PLoS ONE, descriptive and reproducibility values are listed of all parameters.
 


[GUIDELINES]
For instructions on analysis with the accompanying matlabfiles, read the instructions and standard operation procedures (SOPs)
For questions, send an email to info.demonsprotocol@gmail.com or vanrijn@amsterdamumc.nl

[STEPS]
SECTION: Protocols and instructions
1. Measurement and analysis protocol, and instructions on analysis and import files in Matlab.

The Eyelink measurement is created in the Experiment software of SR research and can therfore only be used with an Eyelink (SR Research) eyetracker. These zipped files contain the folders of the developed experiment. 
For other eyetracker, the experiment has to be created as indicated in the measurement protocol .pdf file
  
 
   
  
  
  

SECTION: Protocols and instructions
2. Matlabfiles needed for automated import of data in Matlab (see document 'SOP_importfiles_eyetracking_08022022')
  
  

SECTION: Protocols and instructions
3. Matlabfiles needed for automated analysis of eye moveement data in Matlab (see document 'SOP_Matlab_analysis_08022022')
                                                                                        

SECTION: Protocols and instructions
4. Matlabfiles needed for visualization of eye moveement data in Matlab (see document 'SOP_Visualization_eyemovementdata_08022022')
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
This protocol outlines the analysis used to plot MetaPhlAn taxonomic assignments. Based on methods from the following publication:<br /><br />Hannigan, Geoffrey D., et al. "The Human Skin Double-Stranded DNA Virome: Topographical and Temporal Diversity, Genetic Enrichment, and Dynamic Associations with the Host Microbiome." <em>mBio</em> 6.5 (2015): e01578-15.

[BEFORE_START]
Supplemental information available at:<br /><br /><a href="https://figshare.com/articles/The_Human_Skin_dsDNA_Virome_Topographical_and_Temporal_Diversity_Genetic_Enrichment_and_Dynamic_Associations_with_the_Host_Microbiome/1281248" target="_blank">https://figshare.com/articles/The_Human_Skin_dsDNA_Virome_Topographical_and_Temporal_Diversity_Genetic_Enrichment_and_Dynamic_Associations_with_the_Host_Microbiome/1281248</a>

[GUIDELINES]
<span class="identifier">sessionInfo</span><span class="paren">(</span><span class="paren">)<br /><br />## R version 3.2.0 (2015-04-16) <br />## Platform: x86_64-apple-darwin13.4.0 (64-bit) <br />## Running under: OS X 10.10.4 (Yosemite) <br />## ## locale: <br />## [1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8 <br />## <br />## attached base packages: <br />## [1] stats graphics grDevices utils datasets methods base <br />## <br />## loaded via a namespace (and not attached): <br />## [1] magrittr_1.5    formatR_1.2    tools_3.2.0    htmltools_0.2.6 <br />## [5] yaml_2.1.13    stringi_0.4-1    rmarkdown_0.7    knitr_1.10.5 <br />## [9] stringr_1.0.0    digest_0.6.8    evaluate_0.7</span>

[STEPS]
?. 
?. 
?. 
?. 
?. 
?. 
?.
TITLE: Modelling human neuronal catecholaminergic pigmentation in rodents recapitulates age-related multisystem neurodegenerative deficits
AUTHORS: Ariadna Laguna, Núria Peñuelas, Miquel Vila

[DESCRIPTION]
Methods used in the manuscript entitled "Modelling
human neuronal catecholaminergic pigmentation in rodents recapitulates
age-related multisystem neurodegenerative deficits"

[STEPS]
TITLE: Low Cost LAMP and RT-LAMP
AUTHORS: Tamara Matute, Isaac Núñez, Fernan Federici

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This protocol collection specifies how to carry out LAMP or RT-LAMP (reverse transcription - Loop-mediated isothermal amplification) reactions with homemade reagents and home-brewed enzymes.</div><div class = "text-block">It describes how to prepare all the solutions and buffers to perform the reactions.</div><div class = "text-block"><span>This collection was developed in the context of COVID-19 pandemic to provide access to RT-LAMP tools in accordance with the principles of </span><a href="https://reclone.org/" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">ReClone</span></a><span> community initiative. It is the result of information exchange and reagent sharing with a huge community of collaborators from ReClone, gLAMP and JOGL.   </span></div></div>

[STEPS]
TITLE: Condor CBD Gummies Website Condor CBD Gummies Website Reviews Ingredients Price Scam Alert: Reviews From The Manufacturers
AUTHORS: Condor CBD Gummies Website

[DESCRIPTION]
https://www.outlookindia.com/outlook-spotlight/-scam-alert-condor-cbd-gummies-reviews-website-is-it-endorsed-by-shark-tank-who-own-it--news-206015

[STEPS]
TITLE:     ELISA for quantification of IL-12 in human serum.
AUTHORS: Angel  Justiz-Vaillant

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">Interleukins (IL) are a type of cytokine first thought to be expressed by leukocytes alone but have later been found to be produced by many other body cells. They play essential roles in the activation and differentiation of immune cells, as well as proliferation, maturation, migration, and adhesion. They also have pro-inflammatory and anti-inflammatory properties. The primary function of interleukins is, therefore, to modulate growth, differentiation, and activation during inflammatory and immune responses. Interleukins consist of a large group of proteins that can elicit many reactions in cells and tissues by binding to high-affinity receptors in cell surfaces. </div><div class = "text-block"><span>Monocytes produce IL-12. Its principal targets are T cells. It causes induction of Th1 cells. Besides, it is a potent inducer of interferon gamma production by T lymphocytes and NK cells. </span><a href="https://www.ncbi.nlm.nih.gov/books/NBK499840" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">[1]</span></a></div><div class = "text-block">Reference</div><div class = "text-block"><span>1. Justiz Vaillant AA, Qurie A. Interleukin. In:</span><span style = "font-style:italic;">StatPearls</span><span>. Treasure Island (FL): StatPearls Publishing; June 12, 2019.</span></div></div>

[STEPS]
?. An anti-human IL-12 coating antibody is adsorbed onto microwells by incubation overnight at 4°C.
?. Add 50 µl of human serum. Human IL-12 present in the serum sample binds to antibodies adsorbed to the microwells.
?. The microplate is blocked with 3% non-fat milk-PBS buffer and later wash to remove unbound proteins.
?. Fifty (50) µl of biotin-conjugated anti-human IL-12 antibody is added. The optimal dilution must be investigated.
?. The microplate is rewashed with PBS-Tween 20 buffer, pH 7.4.
?. One hundred µl of streptavidin-HRP conjugate is added and it binds to the biotin-conjugated anti-human IL-12 antibody. The optimal dilution of this conjugate must be investigated.
?. The plate is washed following incubation to remove the unbound Streptavidin-HRP.
?. Add 100 µl of 3,3',5,5'- tetramethylbenzidine (TMB; Sigma-Aldrich) to each well.
?. Incubate the microwells in the dark for 15 min.
?. A colored product is formed in proportion to the quantity of human IL-12 present in the sample or standard.
?. The reaction is terminated by addition of 100 µl 3M H2SO4  and absorbance is measured at 450 nm.
?. A standard curve is made from 7 human IL-12 standard dilutions and the human IL-12 sample concentration is determined.
?. For better results place the microplate on a plate shaker in every incubation.
TITLE: Addition of the adaptor to RNA substrates for 3′ RACE (mapping P ends)
AUTHORS: Matus Valach

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>Simple protocol for mapping 3′-P RNA termini by RT-PCR after the addition of a 3′ adaptor using </span><span style = "font-style:italic;">E. coli</span><span> RtcB RNA ligase.</span></div></div>

[STEPS]
?. Mix the following components (9 μL):  ABC1ComponentAmount [μL]Final concentration2RNA [1 µg]4100 ng/µL35′-hydroxylated 3′ RACE RNA oligo [100 µM]0.44RNase-free water (ddH2O)4.6
ABC1ComponentAmount [μL]Final concentration2RNA [1 µg]4100 ng/µL35′-hydroxylated 3′ RACE RNA oligo [100 µM]0.44RNase-free water (ddH2O)4.6
?. Denature for 2 min at 70 °C, place on ice.
?. Mix the following components (20 µL):  ABC1ComponentAmount [μL]Final concentration2RNA + oligo mix (step 2)9310× RtcB RNA ligase buffer 21×450% PEG-8000410%5GTP [1 mM]20.1 mM6MnCl2 [10 mM]21 mM7EcRtcB RNA ligase [15 µM]10.75 µM
ABC1ComponentAmount [μL]Final concentration2RNA + oligo mix (step 2)9310× RtcB RNA ligase buffer 21×450% PEG-8000410%5GTP [1 mM]20.1 mM6MnCl2 [10 mM]21 mM7EcRtcB RNA ligase [15 µM]10.75 µM
?. Incubate for 120 min at 37 °C.
?. Purify the RNA from the RNA ligase reaction (e.g., trizol extraction or column clean-up).
?. Proceed to RT-PCR. For the RT reaction, use 250-500 ng of the purified adaptor-ligated RNA and a primer reverse complementary to the 3′ adaptor. For the PCR, use an upstream (forward) primer binding to the RNA of interest and a downstream (reverse) primer binding to the 3′ adaptor.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
Human primary T cells are invaluable and feasible model systems to study the characteristics of the human immune cells in various contexts, including but not limited to cancer immunotherapy. Following isolation of T cells from fresh human blood samples, it is possible to culture, expand, and manipulate these cells, which allows extensive investigation for research purposes. Techniques for isolation and handling of T cells are well-established but parts of the protocols can highly vary across different labs. These differences in the protocols are, often, there due to historical reasons and are only supported by anecdotal evidence. We systematically modified basic components of the T cell culturing protocols and collected data on how they altered the final yield. Here, based on these data, we provide practical hints and tips on basic cellular and molecular techniques for handling primary human T cells. We hope that this guide will serve as a reference point to allow evaluate, discuss, and improve current practices in T cell culturing and manipulation.

[STEPS]
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>This is an excellent rhinovirus screening assay which I have used on thousands of sample extracts mostly originating from acutely ill paediatric patients, spanning well over a decade's worth of collection dates.</p>
<p> </p>
<p>I have not confirmed that it can detect every single genotype but I do know that it detects <em>many</em> from each of the three RV species (<em>Human rhinovirus A</em>, <em>Human rhinovirus B</em> and <em>Human rhinovirus C</em>) as well as at least some <em>Human enterovirus</em> genotypes.</p>
<p> </p>
<p>The assay does pick up some enteroviruses due to their genetic similarities in the 5'UTR target region. These can be discriminated using subgenomic sequencing (see  VP42 typing assay protocol), or simply described as "respiratory enteroviruses" since there is no specific-specific vaccine or treatment available anyway. </p>
<p> </p>
<p>The assay was created and first described in 2008 by Xiaoyan Lu and colleagues at the US CDC [Ref 1]. In collaboration with Lu et al., we published a small update that included Lu's modified forward primer design. We routinely use it with this primer (hence the 'mod' text added to the primer name) but without the expensive special base, as outlined in Step 1.  </p>
<p> </p>

[BEFORE_START]
<div>
<div>
<ul>
<li>If using a different brand or model of real-time thermocycler, check the concentration of ROX is adequate.</li>
<li>Method assumes the user is familiar with the thermocycler and software used to run the protocol.</li>
</ul>
</div>
</div>

[STEPS]
?. 
?. 
?. 
?. 
?.