eamag/protocols-with-steps huggingface dataset:
TITLE: Growing overnight culture of OP50 as worm food
AUTHORS: Bonnie  Evans

[DESCRIPTION]
Luria broth (LB) is a nutrient-rich media commonly used to culture bacteria in the lab. LB agar plates are frequently used to isolate individual (clonal) colonies of bacteria carrying a specific plasmid. However, a liquid culture is capable of supporting a higher density of bacteria and is used to grow up sufficient numbers of bacteria necessary to isolate enough plasmid DNA for experimental use. The following protocol is for inoculating an overnight culture of liquid LB with bacteria. 

This protocol is specifically for making a liquid bacterial culture of OP50 and the following parameters are specific for this bacterial strain:

1. Name of the bacterial strain: E.coli (OP50)
2. Growth temperature: 37C
3. Incubation time: 16-18hrs (Overnight)
4. Rpm of the shaking incubator: 200-220 rpm

[STEPS]
5. Obtain 1L LB Broth from the media kitchen

LB Broth contents: 
4gNaCl
4 g Tryptone
2 g Yeast Extract dH2O to 400 mL 
6. Obtain 2 x 2L sterile Erlenmeyer flask from the glassware room
7. In the microbial hood, add 500mL liquid LB to each flask
8. Using a sterile pipette tip, select a single colony from your plate. Drop pipette tip into the flask. 
9. Re-cover the flask with the sterile aluminium foil
10. Incubate the bacterial cultures at the 37C 200rpm for 16-18 hours. 
 

11. After incubation, check for growth, which is characterized by a cloudy haze in the media. 
12. Measure the optical density of the overnight culture. 
 



 

3. Streak OP50 onto LB Agar and grow overnight at 37C.


 


1. Obtain 100mL LB Agar from the media kitchen. Heat in the microwave in 20s bursts until liquid. 


2. Label 2 x 90mm Petri dishes with 'LB Agar' and the date. Using a stripette, dispense 35mL LB agar into each of them. Leave to set on bench. 
4. The next day, check for individual colonies on OP50 plate. Parafilm the plate and store in 4C fridge for up to a month. 
13. Adjust OD600 of the culture to 1.0 using LB. 

Amount of LB (mL) to add = (OD600 x 500mL) - 500mL
14. Aliquot culture into 50mL Falcon tubes and store in 4C fridge.
TITLE: NEBNext End Repair Module E6050
AUTHORS: Isabel Gautreau

[DESCRIPTION]
<div class = "text-blocks"></div>

[STEPS]
?. [Mix the following components in a sterile microfuge tube:]
Fragmented DNA: variableNEBNext End Repair Reaction Buffer (10X): 10 μlNEBNext End Repair Enzyme Mix: 5 μlSterile H20 for a final volume of 100 μl: variableTotal volume: 100 μl
?. Incubate in a thermal cycler for 30 minutes at 20°C.
?. Purify DNA sample on one column.
TITLE: SOP for cDNA synthesis by RT (Reverse Transcription)  Promega kit
AUTHORS: Malu G Tansey

[DESCRIPTION]
SOP for cDNA synthesis by RT (Reverse Transcription)  Promega kit

[STEPS]
SECTION: Protocol:
2. Prepare per sample: 

 

SECTION: Protocol:
3. Heat samples at 70°C for 5 min (prepare mix for step 4).
SECTION: Protocol:
4. Snap freeze samples at +4°C or on ice for a minimum of 5 min.
SECTION: Protocol:
5. Prepare RT mix (vol per samples, calculate for all your samples + 1):
 

SECTION: Protocol:
6. Brief centrifugation.
SECTION: Protocol:
7. RT program
 

SECTION: Protocol:
8. Add H2O to bring final concentration to a desired amount – this will likely depend on the abundance of your targets.
SECTION: Protocol:
9. Once it is done, cDNA can be stored at -20°C
SECTION: Before Begining:
1. Take from the kit: OligodT, RT buffer (5x), MgCl2, dNTP mix, and H2O to thaw- you can keep them on ice or at RT for a little while.
TITLE:         ELISA for quantification of human C2 in serum or plasma.
AUTHORS: Angel  Justiz-Vaillant, Belkis Ferrer-Cosme

[DESCRIPTION]
<div class = "text-blocks"></div>

[STEPS]
?. An anti-human C2 coating antibody is adsorbed onto the microwells by incubation overnight at 4°C with carbonate-bicarbonate buffer.
?. Add 50 µl of human serum or plasma. Human C2 present in the serum or plasma binds to antibodies adsorbed into 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-C2 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-C2 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) into each well.
?. Incubate the microwells in the dark for 20 min.
?. A colored product is formed in proportion to the quantity of C2 present in the sample or standard.
?. The reaction is terminated by addition of 100 µl 3M H2SO4  and the absorbance is measured at 450 nm.
?. A standard curve is made from 7 human C2 standard dilutions and the human C2 sample concentration is determined.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>Quality impact results is the universal first step for all sequencing methods. This protocol is for sequencing results in FASTQ format. FastQC allows for visualization of quality scores. Sequence trimming allows for removal of residual primer sequences and increases overall quality scores. Trimmomatic does both. CutAdapt cuts adapters from sequences. Sickle creates a sliding window for quality and length trimming of FASTQ files.</p>
<p> </p>
<p>Open this protocol inside the virtual machine (details in 'Start Instructions') for easy copy, paste of commands into the command line terminal window.</p>

[BEFORE_START]
<p>Before starting, please visit the ECOGEO website for more information on this 'Introduction to Environmental 'Omics' training series. The site contains a pre-packaged virtual machine that can be downloaded and used to run all of the protocols in this protocols.io collection. In addition to the VM, the website contains video and presentations from our initial 'Intro to Env 'Omics' workshop held at the Univ. of Hawai'i at Manoa on 25-26 Jul 2016.<br /><br />Please email ‘ecogeo-join@earthcube.org’ to join the ECOGEO listserv for future updates.</p>

[STEPS]
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?. 
?.
TITLE: Free floating immunofluorescence protocol on mouse brain sections for tau pathology
AUTHORS: Felicia Suteja, Hongyun Li, YuHong Fu

[DESCRIPTION]
This protocol describes our free-floating  multiplexed immunofluorescent staining protocol to ascertain levels of tau and phospho- tau in mouse tissue from transplanted human iPSC cells carrying different PD related mutations.

[GUIDELINES]
IMPORTANT: perform all antibody incubation steps and steps following in minimal light so as not to bleach signals prior to imaging

[STEPS]
SECTION: Experimental outline
1. Briefly, the mouse brain tissue sections are prepared by washing off the cryoprotectant medium and then antigen retrieval is performed followed by quenching, blocking and primary antibody incubation. Sections are then washed and incubated in the appropriate secondary antibody solution and are then mounted, cover-slipped and sealed.
SECTION: Day 1 - Tissue prep
2. 30 μm mouse brain sections were stored in anti-freeze solution at -20 °C  until required. 

Pour sections into a well insert in a 6-well plate to separate storage solution from section
 Move the well insert to another well containing approximately 6 mL of 1x PBS. Wash at least 6x with 1x PBS for 5 min each on an orbital shaker using low speed at

SECTION: Antigen retrieval
3. Place sections in labelled glass vials containing 6 mL 1x citric buffer (CB) pH6.0.
Place in the steamer (Breville, Model: BFS800BSS) on high for 22 min 98 °C.
Cool down to Room temperature
Place the sections back into its corresponding wells.
1xPBS wash: 2 x 5 min
SECTION: Blocking and primary incubation
4. Incubate in blocking buffer: 120 min Room temperature on shaker 
2. Make primary antibody cocktail in home-made IF buffer
3. Label ceramic plate and place sections in antibody cocktail: 4320 min at 4 °C on the shaker
 - AT8(Ms IgG1), Thermofisher #MN1020             1:200
 - SP70 (rb), Sigma #SAB5500182	                 1:200
 - TH (Ms IgG2b), Thermofisher # TA506549      1:200
SECTION: Day 4 - Secondary antibodies
5. Transfer sections from ceramic plate to well plates
0.1% PBST wash: 3 x10 min
Make secondary cocktails as below in IF blocking buffer
 - Gt @ Ms IgG,  ThermoFisher #A-21206	                          1:250
 - Dn@Rb-AF568, ThermoFisher #A-10042                          1:250
 - Gt@Ms IgG2b-AF647, Thermofisher #A-#A-21242          1:200
- Hoechst 33,342 (1mg/ml stock) Sigma #B2261                 1:1000
4. Place sections and antibody cocktails in a ceramic plate: 120 min Room temperature on the shaker  
5. Place sections back into wells
6. 0.1% PBST wash: 3 x 10 min
7. 1x PBS wash: 3 x 5 min 
SECTION: Mounting and sealing
6. Label slides with corresponding section and antibody information
Place sections in a petri dish with 1xPBS and nudge the sections onto slides with brush
Mount with anti-fade media (DAKO Fluorescence Mounting Medium, Agilent, cat# S302380-2)
Wait for sections to become dry and seal with sealant (Biotium CoverGripTM Coverslip Sealant, cat#23005)
Store in slide box and store in fridge/cool room 5 min
TITLE: DNA extraction, amplification, and sequencing of Ophiothrix (Echinodermata: Ophiuroidea)
AUTHORS: Renata Alitto, Michela Borges, Letícia Dias de Oliveira, Karin Regina Seger

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>DNA extraction, amplification, and sequencing, particularly for the brittel star </span><span style = "font-style:italic;">Ophiothrix.</span></div></div>

[STEPS]
?. [DNA extraction]
Samples of tube feet from the arms or gonads were soaked in two changes of Tris-EDTA (TE) buffer for 30 minutes each. The samples were then macerated in 500 μl of TE buffer with a pestle. A volume of 300 μl of a 10 % Chelex 100 solution (BioRad) was added. The samples were then incubated at 55°C for 60 minutes, boiled for 8 minutes, cooled to room temperature, vortexed for 20 seconds followed by 60 seconds of centrifugation at 14,000 xg. The supernatant was decanted and kept refrigerated until use in polymerase chain reactions (PCR)
?. [Amplification]
A fragment of the mitochondrial 16S gene was PCR-amplified using the forward primer 16S Sofi F (5′-CAGTACTCTGACTGTGCAA-3′) and the reverse primer 16S Sofi R (5′-GGAAACTATGATCCAACATC-3′). A fragment of the mitochondrial cytochrome oxidase subunit 1 (COI) gene was amplified using the forward primer COIf-L (5′-CCTGCAGGAGGGGGAGAYCC-3′) and the reverse primer COIa-H (5′-TGTATAGGCGTCTGGATAGTC-3′). PCR reactions were performed using PuReTaq Ready-To-Go™ PCR Beads for 25 μl reactions (GE Heathcare), complemented with 0.5 to 1 μl of 25mM MgCl2. Alternatively, PCR reactions were composed of 10 mM Tris-HCl, pH 8/1 mM KCl buffer, 1.5 mM MgCl2, and 200 μM each dNTP (Invitrogen). In both cases, 0.4 μM of each primer was used and template DNA concentration (ranging from 25.7 to 917.9 ng/μl) was tested.Thermal cycling consisted of a single step at 94°C for 5 minutes, which was followed by 39 cycles (denaturation at 94°C for 45 seconds, annealing at 45.7°C to 51.3°C for 45 seconds, and extension at 72°C for 60 seconds) and a final extension at 72°C for 3 minutes on a thermal cycler (Eppendorf Mastercycler®).
?. [Purification]
PCR products were separated from excess primers and dNTP using a purification kit (Promega). Purified product was then used as a template for DNA sequencing.
?. [Sequence]
Reactions were done with the BigDye Terminator (Applied Biosystems)Serviço de Sequenciamento de DNA – SSDNA IQUSP with the same primers used in the amplification reaction.
TITLE: Performance comparison of stress hyperglycemia ratio for predicting fatal outcomes in patients with thrombolyzed acute ischemic stroke
AUTHORS: Sarawut Krongsut, Chatchon Kaewkrasaesin

[DESCRIPTION]
Stroke is a prevalent neurological condition and a primary global cause of death, resulting in
approximately 6 million annual fatalities [1].Stroke is the leading cause of death in Thailand, accounting for over 250,000 new cases and 50,000 annual fatalities [2].  Recombinant tissue plasminogen activator (rt-PA) is recommended as a safe and effective treatment [3]. Elevated blood sugar in 40-50%of acute stroke patients may exacerbate ischemic injury through heightened oxidative stress, endothelial dysfunction, and impaired fibrinolysis, resulting in larger infarctions, worse clinical outcomes, and increased mortality rates.[4-5]. Stress hyperglycemia (SH) refers to transient hyperglycemia in the context of illness accompanied by diabetes mellitus (DM) or non-DM. Recently, Roberts et al. [6] introduced the
stress hyperglycemia ratio (SHR) to evaluate SH. Hemoglobin A1c (HbA1c), a stable indicator, was used to assess glycemic management in DM patients over three months. SHR is calculated by dividing the admission glucose concentration by the estimated average glucose concentration derived from HbA1c [7]. Different studies employed the glucose/HbA1c ratio to define SHR, aiming for its practical use in clinical settings [8-10].

Poor outcomes and symptomatic intracerebral hemorrhage (sICH) in acute ischemic stroke (AIS) patients treated with rt-PA were associated with hyperglycemia. According to the American Diabetes Association, patients were classified as DM, newly diagnosed DM, or experiencing transient hyperglycemia during hospitalization. The definition of SH remains unclear, but an abrupt increase in plasma glucose levels above the average blood glucose level serves as a reliable indicator [6,11]. Two types of biological markers for SH, SHR and glycemic gap (GG), have been developed to represent SH [11]. Recently, the SHR, a ratio of plasma glucose level to HbA1c, has emerged as a prognostic biomarker for poor outcomes in AIS patients receiving rt-PA treatment.

Although different SHR equations effectively predicted unfavorable outcomes or critical illness in AIS patients [12], the optimal threshold of SHR for assessing SH and predicting fatal outcomes (in-hospital mortality [IHM], malignant
cerebral edema [MCE], and sICH) has not been definitively confirmed.  Limited data currently exists regarding the
comparative predictive value of various types of SHR, GG, absolute plasma glucose, and HbA1c in predicting fatal outcomes in AIS patients treated with rt-PA. Hence, this study aims to explore the predictive performance, optimal thresholds, and association between these variables in predicting fatal outcomes.



 




 
 

[STEPS]
1. Performance comparison of stress hyperglycemia ratio for predicting fatal outcomes in patients with thrombolyzed acute ischemic stroke
TITLE: Sticker Removal
AUTHORS: Ian N Krout, Tim Sampson

[DESCRIPTION]
The sticker removal test is a form of adhesive removal task assessment that is used by our lab to asses sensory stimuli and motor tasks in mice. Simply, an adhesive substance is placed on the nose of the mouse which is sensed, then removed by the mice using their forepaws1. Difficulties in these tasks are commonly attributed to either sensory perturbations or difficulty in initiating or performing movement2. PD models (Parkin-KO, DJ-1, and Thy1-aSyn) mice show an increase in the time required to remove the adhesive sticker compared to controls2.

[STEPS]
SECTION: Acclimation
1. Bring mice up from vivarium in their home cage to acclimate for at least 1h prior to sticker removal assessment.
SECTION: Assessment
3.     
With one hand use the tweezers to pick up one sticker, allowing it to sit on a single prong with the
adhesive side freely visible.
SECTION: Assessment
4.     
With the free hand grab one mouse and manually restrain with a “scruff technique” immobilizing the head and forepaws.
SECTION: Acclimation
2. Randomize the order of mice for assessment and record this order in your lab notebook
SECTION: Assessment
5. Using the tweezers gently press the adhesive side of the sticker onto the snout of the restrained mouse, gently putting down the tweezers and picking up a stopwatch.
SECTION: Assessment
5.1. Avoid placing the tweezers onto a hard surface or dropping them, as the high pitch noise will disturb the mice.
SECTION: Assessment
6.     
Place the restrained mouse into a clean, empty, cage and allow the mouse to remove the sticker from the snout, using its forepaws. 
SECTION: Assessment
7. Stop the timer once the sticker has been removed completely from the snout and remove the sticker (now on the forepaws) from the mouse. 
SECTION: Assessment
8. Allow the mouse 30 seconds to recover in the clean cage.
SECTION: Assessment
9. Repeat 2x times for a total of 3 replications.
SECTION: Assessment
10. Move on to the next mouse.
SECTION: Assessment
10.1. Clean the cage with 70% EtOH or use a new cage for the assessment when moving onto the next home cage.
SECTION: Assessment
11. Record the adhesive removal time as the average of 3 replications.
TITLE: Lentivirus production
AUTHORS: Itika Saha, F. Ulrich Hartl, Mark S. Hipp

[DESCRIPTION]
This protocol describes the production of lentiviruses to transduce HEK293T cells and has to be performed in a biosafety level 2 laboratory

[STEPS]
SECTION: Lentivirus production
1. Plate ~3.6x106 Lenti-X HEK293T cells (Takara) in 10 cm dish in 10 mL standard DMEM. Cells should be ~80% confluent at the time of transfection. 
 

SECTION: Lentivirus production
2. Next day, remove 5 mL medium and replenish with fresh medium.
SECTION: Lentivirus production
3. Warm up reduced serum medium e.g. Opti-MEM (Gibco) and transfection reagent to room temperature (RT). This protocol was performed with Lipofectamine 3000 transfection reagent (Thermo).
SECTION: Lentivirus production
4. Add 24 µL Lipofectamine 3000 to 600 µL Opti-MEM, mix by vortexing and incubate 5 min at RT.
SECTION: Lentivirus production
5. In another tube, mix 6 µg plasmid containing gene of interest, 5 µg packaging plasmid psPAX2 
(RRID:Addgene_12260), 1 µg envelope plasmid pMD2.G (RRID:Addgene_12259) and 24 µL P3000 reagent (provided by manufacturer along with Lipofectamine 3000 reagent) in 600 µL Opti-MEM, mix by vortexing and incubate 5 min at RT.
SECTION: Lentivirus production
6. Mix contents of both tubes and incubate for 15 min at RT.
SECTION: Lentivirus production
7. Add DNA-lipid complex to cells dropwise.
SECTION: Lentivirus production
8. 2 days later, collect virus-containing medium and centrifuge for 5 min at 1,000 x g.
SECTION: Lentivirus production
9. Collect supernatant in a fresh tube and proceed with concentration.
SECTION: Concentration
11. Incubate overnight or 2 h at 4 °C.
SECTION: Concentration
12. Next day, centrifuge for 45 min at 1,500 x g at 4 °C followed by gently aspirating supernatant.
SECTION: Concentration
13. Resuspend viral pellet in 100 - 1000 µL PBS, aliquot and store at -80 °C until use.
SECTION: Concentration
10. Add Lenti-X concentrator (Takara) to clarified virus-containing medium at 1:4 dilution and mix well by gently inverting tube.
TITLE: Preparation of s/o/w emulsion using SPG membrane
AUTHORS: Daisuke Yasui

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">Solid-in-oil-in-water emulsion of miriplatin is prepared by pushing miriplatin suspension through 20 μm SPG membrane (SPG technology) using a syringe pump.</div><div class = "text-block">0.8w% HCO solution is used as a outer aqueous phase.</div></div>

[STEPS]
?. Prepare  half-saline and add   of Polyoxyethylene hydrogenated castor oil 60 (HCO 60).Use a starler to make the solution.
10 mL
0.08 mg
?. Aspirate the solution to 10mL syringe and connect it to a filter kit with 20μm hydrophilic-coated SPG membrage(SPG Technology Co.,Ltd).Push the whole solution into the filter kit.
?. Aspirate   of Lipiodol (Gurbet, Villepinte, France) in a syringe and add it to the bottle of miriplatin (Dainippon Sumitomo Pharma, Osaka, Japan), thus prepare miriplatin suspension.
3 mL
?. Aspirate miriplatin suspension with a syringe and connect to the filter kit, which was prepared in step 2.First, aspirate air in the system and then set a syringe to a syringe pump.Set stirler beneath the filter kit.Start a syringe pump with a speed of 20ml/hr and start a stirler at the same time with a speed of 300rpm.
?. Increase the speed of stirler by 200ppm every 5 minuites.
TITLE: Bradford Assay with RotiQuant (ROTH)
AUTHORS: iGEM Dusseldorf

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">Assay for quantification of protein by comparing measured Absorbance at 595 nm to bovine serum albumine standard</div></div>

[STEPS]
?. [Standard measurements]
Dilute your BSA stock solution to 200µg/ml in the same buffer used for your solution of interest.
?. [Standard measurements]
Dilute the Roti-Quant 5x reagent at a rate of 2:7,5. For 15 ml solution, this means 4 ml Roti-Quant added to 11 ml of Water.
?. [Standard measurements]
Create 200 µl each of standard concentrations according to the following table:  ABC1Concentration [µg/ml]Volume of stock neededVolume of solution buffer needed200200320201804404016056060140680801207100100100
ABC1Concentration [µg/ml]Volume of stock neededVolume of solution buffer needed200200320201804404016056060140680801207100100100
?. [Standard measurements]
Work in Triplicates. Pipette 50µl of the standards into the 96-Well plate
?. [Standard measurements]
Add 200 µl of the diluted Bradford reagent to each well.
?. [Standard measurements]
Incubate for 5 minutes at room temperature
?. [Standard measurements]
Measure the OD595 for each well. Create a regression line from the averages of each concentrations' measurements
?. [Protein concentration measurements]
Dilute your solution of interest 1:20 or 1:40, depending on expected protein levels.
?. [Protein concentration measurements]
Work in triplicates and use blanks. Pipette 50 µl of each dilution into a well on the 96-Well plate.
?. [Protein concentration measurements]
When working with solutions containing pigments, prepare additional wells to which NO Bradford reagent will be added
?. [Protein concentration measurements]
Add 200 µl of the diluted Bradford reagent to each well.  For the pigment controls, add 200 µl of solution buffer instead. Optimally, use a multi-pipette to reduce the time needed for adding the reagent.
?. [Protein concentration measurements]
Incubate for 5 minutes at room temperature
?. [Protein concentration measurements]
Measure the OD595 for each well.
?. [Protein concentration measurements]
When processing the data, correct the measurements for the blanks. When pigments are present in the solution, correct the OD595 by subtracting the OD595 of the pigment controls from the appropriate measurements.
?. [Protein concentration measurements]
Determine the amount of protein by putting the achieved OD595 of your solution of interest into the regression formula and multiplying by the dilution factor.
TITLE: Testing For Cellulose Degradation
AUTHORS: Jiri Hulcr

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This protocol describes how to test for cellulose degradation using the congo red method.</div><div class = "text-block"><span>This protocol is part of the Bark Beetle Mycobiome (BBM) Research Coordination Network. For more information on the BBM international network: Hulcr J, Barnes I, De Beer ZW, Duong TA, Gazis R, Johnson AJ, Jusino MA, Kasson MT, Li Y, Lynch S, Mayers C, Musvuugwa T, Roets F, Seltmann KC, Six D, Vanderpool D, & Villari C. 2020. Bark beetle mycobiome: collaboratively defined research priorities on a widespread insect-fungus symbiosis. Symbiosis 81: 101–113 </span><a href="https://doi.org/10.1007/s13199-020-00686-9" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">https://doi.org/10.1007/s13199-020-00686-9</span></a><span>.</span></div></div>

[STEPS]
TITLE: Nuclei Isolation from Human Synovium and Infrapatellar Fat Pad for 10x Multiome
AUTHORS: Irene Lorenzo Gomez, Erpei Wang, Merissa Olmer, Martin Lotz

[DESCRIPTION]
This protocol describes isolation of nuclei from fresh-frozen human knee synovium and fat pad for use in Omics analyses, including RNA-sequencing of ATAC-sequencing. Tissue dissociation was the most critical step in optimization which was monitored by nuclei yield, integrity and purity. Methods for quality control are also described.

[STEPS]