The largest database of trusted experimental protocols

Tris acetate edta buffer

Manufactured by Merck Group
Sourced in United States

Tris-acetate EDTA Buffer is a laboratory buffer solution used to maintain a specific pH range in various biochemical and molecular biology applications. It is a commonly used buffer in procedures such as gel electrophoresis, DNA and RNA purification, and enzymatic reactions. The buffer helps to stabilize the pH environment, which is crucial for the proper function and integrity of biomolecules.

Automatically generated - may contain errors

9 protocols using tris acetate edta buffer

1

Agarose Gel Electrophoresis Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols
The PCR products were tested for amplification of specific genes by agarose gel electrophoresis using 2.0% agarose gel in ×1 Tris-acetate EDTA Buffer (Sigma-Aldrich). A total volume of 60 mL of 2.0% agarose (Sigma-Aldrich) was prepared in ×1 Tris-acetate EDTA Buffer and placed in microwave oven until melted. Molten agarose was allowed to cool to about 55°C and ethidium bromide was added to give a final concentration of 0.5 µg/mL. The gel was poured onto electrophoresis through fitted with a comb. The gel was allowed to set on a flat surface for about 15 min. Electrophoresis was placed in an electrophoresis tank filled with ×1 Tris-acetate EDTA Buffer. Samples were prepared on a parafilm by mixing 2 µL of Gel Loading Buffer (Sigma-Aldrich) and 8 µL of PCR products were loaded in parallel with 100 bp ladder (Direct load PCR 100 bp low ladder, Sigma-Aldrich). Electrophoresis was done at 70 volts for 10 min, then at 50 volts for 2 h. Gel was viewed under an ultraviolet transilluminator and photographed with gel documentation system (BIO RAD Gel Doc EZ Images, USA) for future analysis.
+ Open protocol
+ Expand
2

Quantifying Poly(I:C) in Engineered Nanoparticles

Check if the same lab product or an alternative is used in the 5 most similar protocols
To qualitatively determine the amount of poly(I:C) within the ENCPs, samples were loaded in an agarose gel at 1% w/v in Tris Acetate-EDTA buffer (Sigma-Aldrich, MO, USA) before and after the incubation with an excess of heparin for poly(I:C) displacement. Each lane was loaded with 2.5 μg of poly(I:C) and with 1x SYBR®Gold nucleic acid stain (Invitrogen, CA, USA). For the displacement with heparin, 20:1 and 500:1 weight ratios of heparin (Sigma-Aldrich, MO, USA) to poly(I:C) were added for the C12r8 or pArg ENCPs, respectively, and incubated for 30 min at 37°C. Control lanes included a DNA 1 kb ladder (Invitrogen, CA, USA), and free poly(I:C) in the same conditions as the ENCPs. Gels were run for 30 min at 90 V in a Sub-Cell GT cell 96/192 (Bio-Rad Laboratories, CA, USA), evaluated with an UV transilluminator (Molecular Imager® Gel Doc™ XR, Bio-Rad Laboratories, CA, USA) and analyzed with Image Lab™ Software (Bio-Rad Laboratories, CA, USA).
For the release of poly(I:C), ENCPs were incubated in cell culture media, in a 1:1 (v/v) ratio, for 4 or 24 h, prior to been processed as described above. Free poly(I:C) exposed to the same conditions was used as the control.
+ Open protocol
+ Expand
3

Lateral Flow Biosensor for Multiplex Detection

Check if the same lab product or an alternative is used in the 5 most similar protocols
Gold nanoparticles (AuNPs), 9.46 nM 10 nm diameter, were purchased from BBI Solutions (Cardiff, UK). NHS-PEG4-Biotin was purchased from Thermo Fisher Scientific, Inc. (Waltham, MA). C-reactive protein (CRP) was purchased from Innovative Research (Novi, MI). Ampicillin was purchased from Carl Roth GmbH (Karlsruhe, Germany). Mouse Fc fragment (mFc) was purchased from Jackson ImmunoResearch Europe Ltd. (Newmarket, UK). Tris-acetate-EDTA buffer (40 mM Tris, 20 mM acetic acid, 1 mM EDTA), sodium chloride, magnesium chloride, calcium chloride, tris(2-carboxyethyl)phosphine (TCEP), streptavidin and all other reagents where purchased from Sigma-Aldrich (Munich, Germany) and were at least of analytical grade. Lateral Flow Strips were prefabricated by R-Biopharm AG (Darmstadt, Germany), with Streptavidin on the test line and anti-mouse antibody from goat on the control line. Pur-A-Lyzer Dialysis Kits, 10 kDa MWCO, were from Sigma-Aldrich (Munich, Germany). 5,000 MWCO centrifugal concentrators were from Sartorius (Goettingen, Germany). Oligonucleotides were from Integrated DNA Technology (Coraville, IA), sequences are as follows:
α-Amp-short: 5′-SH-(C6H12)-CAC GGC ATG GTG GGC GTC GTG-3′
α-Amp-pol(T): 5′-SH-(C6H12)-TTT TTT TTT TTT TTT CAC GGC ATG GTG GGC GTC GTG-3′
α-CRP: 5′-SH-(C6H12)-ACA CGA TGG GGG GGT ATG ATT TGA TGT GGT TGT TGC ATG ATC GTG G-3′
+ Open protocol
+ Expand
4

Molecular Biology Toolkit: Reagents Inventory

Check if the same lab product or an alternative is used in the 5 most similar protocols
Ethidium bromide (EtBr), ethylene glycol-bis (β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid (EGTA), ethylenediaminetetra acetic acid (EDTA), isopropyl-β-D-thiogalactoside (IPTG), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) coelenterazine, carbenicillin, CaCl2, Carbonyl cyanide m-chloro phenylhydrazone (CCCP), Calmidazolium (CDZ), Verapamil, coelenterazine, Trifluoperazine (TPZ), Chlorpromazine (CPZ), Phosphate Buffer Solution (PBS), Triton X-100, Tris-Acetate EDTA buffer (TAE), and aequorin oligonucleotides were all purchased from Sigma-Aldrich (St. Louis, MO, United States). Taq polymerase master mix for PCR (Promega, Madison, WI, United States), agarose molecular grade and 100 bp molecular ruler were obtained from Bio-Rad, Hercules, CA, United States). Kanamycin, Gentamicin, Streptomycin, Vancomycin, Ciprofloxacin, and Erythromycin (St. Louis, MO, United States). BamHI and HindIII (Rowley, MA, United States), T4 ligase (Rowley, MA, United States).
+ Open protocol
+ Expand
5

Amplification of spa Gene Region

Check if the same lab product or an alternative is used in the 5 most similar protocols
Published primers were used to amplify the X region of the spa gene [13 (link)]. Each 25 µl PCR reaction mix included 10.5 µl of sterile nuclease-free water, 12.5 µl of Dream Taq mix (Thermo Fisher Scientific,Waltham, Massachusetts, USA) and 0.5 µl (10 pmol) of both the forward and reverse primers with 1 µl (approximately 50 ng) of template DNA. Cycling was performed in a GeneAmp 9700 PCR System (Applied Biosystems, Foster City, California, USA) with the following conditions: initial denaturation at 95 °C for 5 min, and 35 cycles of denaturation at 95 °C for 45 s, primer annealing at 60 °C for 30 s, extension at 72 °C for 90 s followed by a final extension at 72 °C for 10 min. Amplification products were resolved on a 1.5 % agarose in 1× Tris Acetate EDTA buffer (Sigma-Aldrich, St Louis, Missouri, USA) for 60 min at 95V and visualized with EZ Vision DNA stain (Amresco, Inc., Cleveland, Ohio, USA) using a UV transilluminator (UVP LLC, Upland, California, USA).
+ Open protocol
+ Expand
6

RFLP-based Genotypic Classification of PsHV

Check if the same lab product or an alternative is used in the 5 most similar protocols
The DNA samples extracted from the PsHV isolates and from the reference virus (KS 144/79) were analyzed by RFLP for genotypic classification using the restriction enzyme PstI (20,000 U/mL, New England BioLabs, Inc., Ipswich, MA, USA), as recommended by the manufacturer.
The resulting DNA fragments were separated by electrophoresis on a 0.5% agarose gel in 1× Tris–acetate–EDTA buffer (Sigma–Aldrich, St. Louis, MO, USA) containing 5 μL of ethidium bromide (10 mg/mL) (Life Technologies, Grand Island, NE, USA) at room temperature under constant voltage of 20 V for about 15 h. The BenchTop 1 kb DNA Ladder (Promega, Madison, WI, USA) was used as a DNA size marker. The gels were observed at 254 nm using a MacroVue UV transilluminator (GE Healthcare, Cleveland, OH, USA) and photographed.
+ Open protocol
+ Expand
7

PCR Screening of B. cereus Virulence Factors

Check if the same lab product or an alternative is used in the 5 most similar protocols
The PCR screening of the genetic determinants of the main putative virulence factors involved in the B. cereus gastrointestinal toxi-infections was performed using the GoTaq® Green Master Mix (Promega, Fitchburg, MA, USA). The different primers used are shown in Table 2. Thermal cycling parameters were 5 min at 94 °C, then 32 cycles of 1 min at 94 °C, 1 min at the annealing temperature (depending on the primer pair, see Table 2) and 1 min at 68 °C, and finally 10 min at 68 °C. The success of amplification was confirmed by gel electrophoresis at 100 V for 28 min on 0.8% agarose gel (Santa Cruz Biotechnology, Dallas, TX, USA) in Tris-Acetate-EDTA buffer (Sigma-Aldrich, St. Louis, MO, USA).
For several B. cereus strains, the presence of Nhe and HBL toxins in the supernatant was also assessed using the immunodetection kit Duopath Cereus Enterotoxins (Merck, Darmstadt, Germany).
+ Open protocol
+ Expand
8

Molecular Detection of MRSA Isolates

Check if the same lab product or an alternative is used in the 5 most similar protocols
Definite identification of MRSA isolates was dependent on the detection of their characteristic mecA gene [15 (link)]. Briefly, genomic DNA was extracted using GeneJET Genomic DNA Purification Kit according to the manufacturer’s instructions (Thermo Fisher Scientific Inc., Vilnius, Lithuania). The mecA gene was amplified, by PCR technique, using a Thermal cycler (Biometra UNO–Thermoblock, Germany) in presence of 1 µL from each particular 10 pmol primer (Table 1), 12.5 µL Cosmo PCR Red Master Mix (Willowfort, Birmingham, UK), 1 µL of 50 ng genomic DNA, and nuclease-free double distilled water to final reaction volume of 25 µL. The mecA gene amplification cycling program includes 1 cycle of initial denaturation at 94 °C for 4 min (min) followed by 35 cycles of denaturation at 94 °C for 1 min, annealing at 55 °C for 1 min, and elongation at 72 °C for 1 min, and lastly 1 cycle of final elongation at 72 °C for 10 min. The obtained PCR products were loaded into 1.5% agarose gel containing 5 µL ethidium bromide (0.05 mg/mL) followed by electrophoresis at 100 V in Tris-acetate EDTA buffer (Sigma Aldrich, Hamburg, Germany). The UV transilluminator (HERMLE Labortechnik GmbH, Wehingen, Germany) was used to observe the mecA gene-specific bands and determine their molecular size with the aid of a 100 bp DNA ladder (Geneaid Biotech Lt., New Taipei City, Taiwan) as a marker [16 (link)].
+ Open protocol
+ Expand
9

Rapid DNA Extraction from Zebrafish Embryos

Check if the same lab product or an alternative is used in the 5 most similar protocols
Larval genomic DNA was prepared using the HotSHOT protocol (60 (link)). Briefly, single injected 1 dpf embryos were dissociated in PCR tubes containing 10 μl of 50 mM NaOH at 95°C for 20 min. After cooling down, 1 μl Tris–HCl, pH 7.5, was added and the preparation was mixed. 1 μl of this mixture was used as a genomic DNA template in PCRs using the primers listed in Table S6. Amplicons were analyzed on 2.5% agarose gels prepared with 1x Tris–acetate–EDTA buffer (Sigma-Aldrich).
+ Open protocol
+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required

Sign up now

Revolutionizing how scientists
search and build protocols!