eamag/protocols-with-steps huggingface dataset:
TITLE: CAMbank: cfDNA BCT Field Processing v1
AUTHORS: Eliah G Overbey, Krista A Ryon, jak, chm2042

[DESCRIPTION]
Field processing of cfDNA BCTs for the Cornell Aerospace Medicine Biobank (CAMbank).

Instructions for preserving: plasma and RBC Pellets.

[STEPS]
SECTION: Perform Venipuncture
1. After venipuncture, invert the tubes gently 8 to 10 times to fully mix tube anticoagulant with blood sample.

Store the tube upright at room temperature until centrifugation.

Note: The tube stabilizes cell-free DNA for up to 14 days at 6 °C to 37 °C and CTCs for up to 7 days at 15 °C to 30 °C.
SECTION: Centrifuge Settings: Plasma Separation
2. Note: A swing bucket centrifuge is required. 

Set centrifuge:
acceleration: 9
deceleration: 9
temperature: RT
duration: 20 minutes
speed: 300xg
SECTION: Centrifuge Settings: Plasma Separation
3. Place the cfDNA BCTs in the centrifuge. 
Place a protective cover over the swing buckets in case of tube leakage. 
Start the centrifuge. 
Stand by the centrifuge until the centrifuge reaches max speed. Listen for signs of imbalance or compromised tube integrity. 
SECTION: Separate Plasma
4. Carefully remove cfDNA BCTs from the centrifuge and inspect for separation of red blood vs plasma layers. 

Transfer cfDNA BCTs to a sanitized laminar flow hood. 
SECTION: Separate Plasma
5. Remove the upper plasma layer and transfer to a new 15mL conical tube. 
Cap the conical tubes and return to the centrifuge. 
SECTION: Aliquot Plasma
9. Carefully remove conical tubes from centrifuge. 
Transfer conical tubes to a sanitized laminar flow hood. 
SECTION: Aliquot Plasma
10. Using a P1000, carefully aliquot plasma into 2D barcoded tubes at 500µL each. 

Return 2D tubes with plasma to the rack and place in the -80C freezer.
SECTION: Centrifuge Settings: Plasma Purification
6. Note: A swing bucket centrifuge is required. 

Set centrifuge:
acceleration: 9
deceleration: 9
temperature: RT
duration: 10 minutes
speed: 5000xg
SECTION: Centrifuge Settings: Plasma Purification
7. Place the conical tubes in the centrifuge. 
Place a protective cover over the swing buckets in case of tube leakage. 
Start the centrifuge. 
Stand by the centrifuge until the centrifuge reaches max speed. Listen for signs of imbalance or compromised tube integrity. 
SECTION: Freeze Red Blood Cell Pellets
8. Recap and freeze the cfDNA tubes to preserve DNA in the red blood cell pellet
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>Exclusion criteria: lower limb varices, history of leg surgery, trauma, infection, or thromboembolic events.</p>
<p> </p>
<p>Ultrasound examinations are performed in B-mode technique, equipped with a linear broadband (7.5-14-MHz) transducer. The right femoral vein and the inguinal ligament are examined in a supine position. The inguinal ligament is identified along its longitudinal axis. The measurements levels are identified as follow:</p>
<p>-    proximal level: 20 mm caudally to the inguinal ligament identified along the femoral artery imaged with US</p>
<p>-    distal level: 20 mm caudally to the inguinal crease visually identified and along the femoral artery imaged with US</p>
<p>The cross-sectional area (CSA) of the right femoral vein is examined at both levels in transverse plane and is measured automatically after manual tracing of the vessel on a static image in three leg positions: abduction, abduction + external rotation, abduction + external rotation + 90º knee flexion/frog-leg. Attention must be taken to avoid any compression or displacement of the examined vein.</p>

[STEPS]
TITLE: test_CB
AUTHORS: Christine Briggs

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This is a test.</div></div>

[STEPS]
?. [old version. This is empty. I'll fork someone else's protocol here.]
This is a link to  the 10x sequencing user guides at https://support.10xgenomics.com/single-cell-gene-expression/library-prep/doc/user-guide-chromium-single-cell-3-reagent-kits-user-guide-v2-chemistry
TITLE: GM Covid-19 saliva test (v1)
AUTHORS:  

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">GM Covid-19 saliva test is a RT-PCR test intended for qualitative detection of nucleic acids from SARS-CoV-2 in saliva specimen. The test is used for screening purpose under class of "research used only" (RUO). The test is not for diagnostic purpose. Positive result would be referred to an official clinical test with appropriate certification (i.e CLIA) required for clinical diagnosis.</div></div>

[STEPS]
?. [Setup reaction]
Prepare PCR master mixSqueeze Bottle A to add all of its liquid into Bottle B; Close lid of Bottle B and shake to mix; Squeeze Bottle B to load one droplet into each PCR tube to setup a 96-well PCR plate.
?. [Setup reaction]
Saliva-PrepSaliva is self-collected into any sterile tubes with screw cap and wide opening (Sanitize hands before and after saliva collection); Use disposable transfer pipette with fixed volume to pipett  saliva into a Saliva-Prep tube, tightly screw cap, and mix thoroughly;Heat the Saliva-Prep tube at  for   by using a dry-bath or equivalent;Put the tube on ice for  , then at  on GM-rack for
70 °C
0 Room temperature
?. [RT-PCR]
Load sampleUse disposable GM-stick to slowly insert into Saliva-prep tube and slowly pull it out, then directly dip it into a PCR tube. Close PCR tube. Repeat this step for each sample.Pulse spin the PCR strip/plate to bring the liquid to bottom.
?. [RT-PCR]
RT-PCRPut the PCR strip/plate into a thermal cycler, choose and start the program "GM Covid19 Test".Note: The test is suitable for conventioinal thermal cylers. BenchMark's TC32 and Thermofisher's Veriti thermocycler have been tested.
?. Check color Check color of PCR reactions while keeping PCR tube closed. There are two ways to check color of PCR reactions:
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
<p>This is the hands-on interactive component to the ECOGEO Workshop Module on Binning.</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>
<p> </p>
<p>This tutorial is tailored for <a href="http://anvio.org" target="_blank">Anvi’o</a> v2.0.1 (installed on the VM). The purpose of this tutorial is to demonstrate some of the anvi’o capabilities using the Infant Gut dataset, which was generated, analyzed, and published by <a href="http://www.ncbi.nlm.nih.gov/pubmed/22936250" target="_blank">Sharon et al. (2013)</a>, and was re-analyzed in the <a href="https://peerj.com/articles/1319/" target="_blank">anvi’o methods paper</a>.</p>
<p> </p>
<p>The tutorial used during the actual workshop is no longer active. A very similar tutorial is available online (<a href="http://merenlab.org/tutorials/infant-gut/" target="_blank">http://merenlab.org/tutorials/infant-gut/</a>) and should be used to explore the entire hands-on demonstration in the video, as well as reading up on Meren's $0.02!</p>

[STEPS]
?. 
?. 
?. 
?. 
?. 
?.
TITLE: No Title
AUTHORS: 

[DESCRIPTION]
Protocol for blunting ends by 3' overhang removal and fill-in of 3' recessed (5' overhang) ends using T4 DNA Polymerase.

[GUIDELINES]
CAUTION: Elevated temperatures, excessive amounts of enzyme, failure to supplement with dNTPs or long reaction times will result in recessed ends due to the 3&acute; &rarr; 5&acute; exonuclease activity of the enzyme.&nbsp;<br /><br />* T4 DNA Polymerase can be used in&nbsp;<a href='https://www.neb.com/products/b7201-nebuffer-1-1' target='_blank'>NEBuffers 1.1</a>,&nbsp;<a href='https://www.neb.com/products/b7202-nebuffer-2-1'>2.1</a>, and&nbsp;<a href='https://www.neb.com/products/b7204-cutsmart-buffer' target='_blank'>CutSmart Buffer</a>&nbsp;as well as&nbsp;<a href='https://www.neb.com/products/b7001-nebuffer-1' target='_blank'>NEBuffers 1</a>,&nbsp;<a href='https://www.neb.com/products/b7002-nebuffer-2'>2</a>, and&nbsp;<a href='https://www.neb.com/products/b7004-nebuffer-4' target='_blank'>4</a>&nbsp;and&nbsp;<a href='https://www.neb.com/products/b0202-t4-dna-ligase-reaction-buffer' target='_blank'>T4 DNA Ligase Reaction Buffer</a>. Optimal activity is observed in&nbsp;<a href='https://www.neb.com/products/b7202-nebuffer-2-1' target='_blank'>NEBuffer 2.1</a>. BSA supplementation is recommended when using a buffer that does not already contain BSA.&nbsp;<br /><br />References:<br /><br />1. Tabor, S. and Struhl, K. (1989). DNA-Dependent DNA Polymerases. In F. M. Ausebel, R. Brent, R. E. Kingston, D. D. Moore, J. G. Seidman, J. A. Smith and K. Struhl(Ed.), Current Protocols in Molecular Biology. 3.5.10-3.5.12. New York: John Wiley &amp; Sons, Inc.<br /><br />2. Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989). Molecular Cloning: A Laboratory Manual. (2nd ed.), 5.44-5.47. Cold Spring Harbor: Cold Spring Harbor Laboratory Press.

[STEPS]
?. 
?. 
?. 
?. 
?.
TITLE: Patient-Centered Education Bundle on Administration of Venous Thromboembolism Prevention
AUTHORS: Oluwafemi P. Owodunni1, Elliott R. Haut, Dauryne L. Shaffer, Deborah B. Hobson, Jiangxia Wang, Gayane Yenokyan, Peggy S. Kraus, Jonathan K. Aboagye, Katherine L. Florecki, Kristen L.W. Webster, Christine G. Holzmueller, Michael B. Streiff, Brandyn D. Lau

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">Venous thromboembolism (VTE), comprised of deep vein thrombosis (DVT) and/or pulmonary embolism (PE), affects 350,000-600,000 individuals in the United States annually. More than 100,000 people die each year in the United States as a result of PE. </div><div class = "text-block">Numerous studies have shown that VTE prophylaxis is vastly underutilized in hospitals and the Agency for Healthcare Research and Quality (AHRQ) has listed strategies to improve VTE prevention on its top ten list for patient safety practices. Consequently, numerous interventions have been implemented to improve prescription of VTE prophylaxis with the implicit assumption that medications prescribed for hospitalized patients will always be administered. </div><div class = "text-block">There are deficits in the administration of Prescribed Venous Thromboembolism Prophylaxis. Although the benefit of VTE prophylaxis is well established, many hospitalized patients do not receive adequate VTE prophylaxis. The rationale for the Patient-centered Education Bundle is driven by the high frequency of patient refusal of VTE prophylaxis and significant knowledge gaps regarding VTE, there is an urgent need to educate patients and families on the importance and benefits of compliance with VTE prophylaxis. As part of a study funded by the Patient-Centered Outcomes Research Institute (PCORI), we first educated nurses on the harms of VTE and the benefits of prophylaxis, showing that this first step intervention improved VTE prophylaxis administration by approximately 10%. The patient-centered education bundle will be delivered as an in-person, 1-on-1 discussion session with a nurse educator. Supporting education materials include a 2-page education sheet and an educational video. </div></div>

[STEPS]
TITLE: OD and GFP Plate Reader Assay (72 h Measurement)
AUTHORS: Alba Balletbó

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span>Fluorescence measurements of Optical Density (OD) and Green fluorescent protein (GFP) in </span><span style = "font-style:italic;">Escherichia coli</span><span style = "font-style:italic;">.</span></div></div>

[STEPS]
?. [Media Preparation]
Prepare the following media and autoclave them according to standard procedures. M9TG Media (Without carbon source):  AB1ReagentAmmount to add for 50 mL2M9 Salts1X3Tryptone0.5 gM9TG Media (0.5% Glycerol):  AB1ReagentAmmount to add for 50 mL2M9 Salts1X3Tryptone0.5 g4Glycerol0.25 gM9TG Media (2% Glycerol):  AB1ReagentAmmount to add for 50 mL2M9 Salts1X3Tryptone0.5 g4Glycerol1 gM9TG Media (0.5% Succinate and 1mM Leucine):  AB1ReagentAmmount to add for 50 mL2M9 Salts1X3Tryptone0.5 g4Succinate0.25 g5Leucine6.56 mg
AB1ReagentAmmount to add for 50 mL2M9 Salts1X3Tryptone0.5 g
AB1ReagentAmmount to add for 50 mL2M9 Salts1X3Tryptone0.5 g4Glycerol0.25 g
AB1ReagentAmmount to add for 50 mL2M9 Salts1X3Tryptone0.5 g4Glycerol1 g
AB1ReagentAmmount to add for 50 mL2M9 Salts1X3Tryptone0.5 g4Succinate0.25 g5Leucine6.56 mg
?. [Plate Reader Protocol]
Set the following variables for the BioTek Synergy MX Microplate Reader:-    Temperature: Setpoint 37°CPreheat before moving to next step. Start Kinetic [Run 72:00:00, Interval 0:04:20]-    Shake: Medium for 0:05 (MM:SS)
-    Read OD600:        Absorbance Endpoint of full plate        Wavelengths: 600        Read Speed: Normal-    Read GFP:     Fluorescence Endpoint of full plate      -   Filter Set:         Excitation: 488/9,0         Emission: 510/9,0         Optics: Bottom         Gain: 100         Read Speed: Normal
?. [Bacteria and Plate Reader Preparation ]
Grow desired bacteria overnight in a 50 ml falcon tub, containing  M9TG media and grown cultures @, 250 RPM overnight.
10 ml
37 °C
Luria-Bertani media is also possible to use, but cells must be washed with PBS buffer prior to plate reader experiment because of the high amount of fluorescence from the yeast extract:After overnight incubation, spin cells down at 4700 x G, for 5 min, discard the supernatant and resuspend in 10 ml PBS. Repeat prior steps and resuspension again in PBS.
?. [Bacteria and Plate Reader Preparation ]
Load    of the corresponding buffer in the wells of the 96-well plate and add  of overnight culture.
190 µl
10 µl
The amount of buffer and overnight culture will depend on the OD after overnight bacterial cultivation. The amounts present in this protocol where determined by an OD ~0.3.
When cell have been grown overnight in LB media, washing steps are require before continuing! (See step 3).
?. [Plate Reader Protocol]
Start plate reader protocol.
?. [Data Analysis]
Retrieve the data from the computer, correct for the measurements for the OD600 and subtract the autofluorescence from the control sample of all the other samples.
TITLE: Dopamine Activation in the vmPFC: Impact on Impulsive Choice in Social Contexts and Neural Connections
AUTHORS: Spencer Lee

[DESCRIPTION]
This study protocol details the experimental procedures aimed at assessing the effects of dopamine activation in the vmPFC on impulsive behavior and social interaction. Experiment 1 will focus on fMRI responses. Experiment 2 will assess behavior using the Go/No-Go Task and Social Interaction Task. Experiment 3 will evaluate the longitudinal impact of vmPFC dopamine activation on both fMRI outcomes and behavioral tasks.

[STEPS]
1. The first experiment aims to determine the effect of dopamine activation in the vmPFC. Long-Evans rats will be used for this study. The experimental group of rats will undergo a procedure where dopamine in the vmPFC is activated by injecting an amphetamine solution. Both the control group and experimental group will undergo whole-brain fMRI scans. Activation patterns will be compared using contact ICA and dual regression methods.
2. The second experiment will aim to determine whether dopamine activation in the vmPFC affects impulsive action and social interaction. For this test, two behavioral experiments will be conducted. The Go/No-Go task will involve multiple FR schedules (F3, F5, etc.), with grain/fat pellets used as rewards. In the social interaction tasks, there will be two major components: assessing social behavior in a 3 chamber cage and introducing a stranger rat in a circular wire cage placed in the left or right chamber of the cage. Another experiment will assess behavior towards intruders. Following the experiments, the rats will undergo fMRI imaging. The behavior from the Go/No-Go task and the social interaction task will be compared, along with their fMRI data. Additionally, the fMRI data between the two behavioral tasks will be compared to explore the neurological relationship between impulsive action and social interaction.
3. The last experiment aimed to investigate the longitudinal effects of dopamine activation in the vmPFC. Rats underwent the same procedure as in experiments 1 and 2, with regular activation. Subsequently, they underwent a specific period without activation. After this period, activation levels were assessed using fMRI imaging, and behavioral tasks were conducted.
TITLE: Preparation of  polygalacturonic acid (PGA) to study virulence in Erwinia/Pectobacterium
AUTHORS: Pol Nadal Jimenez, Rita S. Valente

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This medium is used for the growth of Erwinia/Pectobacterium species to ensure that virulence genes are turned ON. These bacteria turn on virulence (plant cell-wall degrading enzymes = PCWDEs) in response to various environmental signals. One of these signals is of plant origin, and it is thought to allow the bacterium to determine whether it has located a potential plant host. These plant signals are sensed by the bacterial sensor/regulator KdgR.</div></div>

[STEPS]
?. [2% PGA stock solution preparation]
Weight  of PGA (Sigma, P3850) using a precision scale.
0.2 g
?. [2% PGA stock solution preparation]
Transfer the powder solution to a   (low-thermal-expansion borosilicate glass) beaker.
100 mL
?. [2% PGA stock solution preparation]
Add   of demineralised water and a (microwave resistant) magnet.
10 mL
?. [2% PGA stock solution preparation]
Place the beaker containing the PGA, the water, and the magnet into a microwave and turn the microwave ON at the maximum power.
?. [2% PGA stock solution preparation]
Stare at the beaker while it's being heated and stop the microwave every time that the solution starts bubbling (be careful, this will happen very sudden and it's easy that you loose the whole mixture if the content overflows the beaker and you don't stop it at the right time).
?. [2% PGA stock solution preparation]
Using some heat-protective gloves remove the beaker form the microwave and place it on a magnetic stirrer. Let the sample cool down while it is being mixed by the magnet.
?. [2% PGA stock solution preparation]
Repeat the process multiple times till the PGA is fully solved in water. The solution will be transparent with a light yellow coloration.
?. [2% PGA stock solution preparation]
Place some sterile aluminium foil on the top of the beaker and keep the solution at room temperature. This solution tends to go off easily, as it has not been properly sterilised. Use it on the same day.
?. [2% PGA stock solution preparation]
Use PGA at a final concentration of  (1 part of PGA stock solution + 4 parts of growth medium)
?.
TITLE: Preparation of BG-11 liquid media 
AUTHORS: Ashwinuday  

[DESCRIPTION]
This is for making liquid media of BG-11.

[STEPS]
1. Take the medium in powdered form and mix it with doubly distilled water to just 1/10 th of the total volume.
2. Add a common stock of micronutrients made in ddH2O and mix well.
3. Make up the volume to the total and mixed well.
4. Adjust the pH on a calibrated pH meter using 1N HCl and /or 1N NaOH. The media was mixed using a magnetic stirrer continuously to homogenize the pH and to prevent precipitation of certain constituents.
5. This mix was autoclaved.
TITLE: cDNA synthesis using SuperScript™ IV
AUTHORS: Roey Angel, Eva Petrova

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">The following protocol is intended as a downstream application for our </div><div class = "text-block"><a href="https://www.protocols.io/view/purification-of-rna-from-a-dna-rna-extract-qdqds5w" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">Purification of RNA from a DNA/RNA Extract</span></a><span> protocol. This protocol describes how to synthesise a first-strand non-specific complementary DNA (cDNA) from a purified RNA extract using </span><a href="https://www.thermofisher.com/cz/en/home/life-science/pcr/reverse-transcription/superscript-iv-reverse-transcriptase.html?gclid=Cj0KCQjw7qn1BRDqARIsAKMbHDa8cd_fkNIkuqyKBkm2FBzP6o0VMxIl4QDHqRZ8QeEk_N1htBuNciwaAlMBEALw_wcB&ef_id=Cj0KCQjw7qn1BRDqARIsAKMbHDa8cd_fkNIkuqyKBkm2FBzP6o0VMxIl4QDHqRZ8QeEk_N1htBuNciwaAlMBEALw_wcB:G:s&s_kwcid=AL!3652!3!285318516879!e!!g!!superscript%20iv?cid=bid_mol_pch_r01_co_cp1358_pjt0000_bid00000_0se_gaw_bt_pur_con" style = "text-decoration:underline;color:blue;cursor:pointer;"><span style = ":;">SuperScript IV Reverse Transcriptase</span></a><span>. The second strand synthesis is usually not required for most downstream applications. This protocol is a simplified and condensed version of the full protocol provided by the manufacturer.</span></div></div>

[STEPS]
?. [Primer annealing]
Prepare the following mixture in a PCR tube:to  ( - ; usually for soil extract)or  a gene-specific primer ()
1 µl
[purified RNA]
10 pg
5 µg
200 ng
[random hexamers (50 µM)]
2
[RNase free water]
?. [Primer annealing]
Mix gently and spin down the solution.
?. [Primer annealing]
Incubate the mixture at for  in a thermocycler and chill  (or in the cycler at >) for at least .
65 °C
on ice
4 °C
?. [Reverse transcription]
Prepare the following mixture and add to each tube:*  *  Optional
[5x Reaction buffer]
[dNTP mix, 10 mM]
[0.1M DTT]
[RNaseOUT™ (40 U/µl)]
[BSA (20 µg/µl)]
[SuperScript™ IV RT (200 units/μl)]
?. [Reverse transcription]
Incubate the mixture in a thermocycler at for  (only if using random hexamers, skip if using a specific primer) followed by  for  to  and  for . Chill .
23 °C
50 °C
80 °C
on ice
?. [Reverse transcription]
For PCR templates > 1kb remove the RNA by adding  (2 units) of E. coli RNase H and incubate atfor .
1 µl
37 °C
?. [Optional: Second strand synthesis]
Prepare the following mixture and add to each tube:
[DNA Polymerase I reaction buffer       ]
[DNA Polymerase I]
[RNase H]
[RNase-free water]
[Template cDNA]
?. [Optional: Second strand synthesis]
Incubate for at  for  followed by  at   for deactivation.
15 °C
75 °C
?. [Optional: Second strand synthesis]
3. Purify the reaction through phenol/chloroform purification followed by ethanol precipitation or using a PCR purification kit.
TITLE: General Taq PCR Master Mix -- CHEM 384/584
AUTHORS: Ken   Christensen

[DESCRIPTION]
2X PCR Master Mixes are convenient to use as they include all the necessary PCR components except the template and primers. Most PCR Master Mixes also include agarose gel running dyes and a density reagent that allows direct loading of PCR products on an agarose gel for electrophoresis. The master mix format simplifies workflows and sample handling; simply add primers, template, and water and then begin PCR. 

[GUIDELINES]
Storage: -20℃ for long-term storage. 4℃ for short-term storage (up to 3 months). (Note) If used frequently, store at 4℃ ; the activity of the Master Mix may decrease with repeated freezing and thawing. Gently mix well before use and centrifuge briefly.

Application: 
・DNA amplification by PCR 
・Colony PCR

PCR Products: Since most PCR products amplified Taq PCR Master Mix have an A overhang added at 3’-termini, the obtained PCR product can be used directly for cloning into a TA cloning vector. Additionally, it is possible to clone the product in a blunt-end vector after blunting and phosphorylation. 

Dye Migration during Electrophoresis: Check individual manufacturers information on the migration of the included running dyes.

[STEPS]
SECTION: Setup Reaction
1. To a 25 µL aliquot of a 2X Taq PCR Master Mix (e.g. TaqDog, or Sapphire Amp), add template (10-20 µl cleared lysate for colony PCR or  20-50 ng of purified DNA for typical PCR), forward and reverse primers to a final concentration of 200 nanomolar (nM). Adjust final volume to 50 µL with nuclease free water or autoclaved water.


 


SECTION: Run Reaction
2. followed by 30 cycles of 98°C, 5 sec; 55°C, 5 sec; and 72°C, 40 sec.