ROS indicator probes APF and HPF, plasmid DNA (pBR322), ethidium bromide, ultrapure 10X TAE buffer and 1 kb plus DNA ladder were purchased from Invitrogen (Life Technologies, Carlsbad, CA). Sodium hypochlorite, silibinin, quercetin, L-methionine, agarose (ultrapure) and calf thymus DNA were purchased from Sigma-Aldrich (St. Louis, MO). Hypochlorite concentration was documented spectrophotometrically in 10 mM NaOH at pH 12 using molar coefficient at 292 nm (ε292 = 350 M−1 cm−1). Dulbecco’s phosphate buffered saline (DPBS ×1, 21-031-CV) without calcium and magnesium was purchased from Mediatech Inc. (Manassas, VA). Commercially available SDG (com) was purchased from Chromadex (Irvine, CA) and synthetic SDGs (SDG (S,S) and SDG (R,R)) were synthesized by our group [39 (link)].
Ultrapure agarose
Manufactured by Merck Group
Sourced in United States, Czechia
Ultrapure agarose is a high-quality laboratory product used for various applications in molecular biology and biochemistry. It is a polysaccharide derived from red seaweed and is commonly used as a gelling agent for the preparation of agarose gels, which are essential for techniques such as DNA electrophoresis, protein separation, and immunoassays.
Automatically generated - may contain errors
Lab products found in correlation
1 kb plus dna ladder, by Thermo Fisher Scientific (2 mentions)
Pbr322 plasmid dna, by Thermo Fisher Scientific (2 mentions)
Calf thymus dna, by Merck Group (2 mentions)
Ethidium bromide, by Thermo Fisher Scientific (2 mentions)
Quercetin, by Merck Group (2 mentions)
Sodium hypochlorite, by Merck Group (2 mentions)
Apf and hpf, by Thermo Fisher Scientific (2 mentions)
Ultrapure 10x tae buffer, by Thermo Fisher Scientific (2 mentions)
L methionine, by Merck Group (2 mentions)
Silibinin, by Merck Group (2 mentions)
Tyrosinase from mushroom, by Merck Group (1 mentions)
Dopamine, by Merck Group (1 mentions)
Triton x 100, by Merck Group (1 mentions)
Trypan blue, by Thermo Fisher Scientific (1 mentions)
Ethidium bromide, by Merck Group (1 mentions)
Trizma base, by Merck Group (1 mentions)
Sodium sarcosinate, by Merck Group (1 mentions)
Low melting temperature agarose, by Merck Group (1 mentions)
Phenol, by Merck Group (1 mentions)
Chloroform, by Merck Group (1 mentions)
6 protocols using ultrapure agarose
1
Oxidative DNA Damage Quantification
2
Oxidative DNA Damage Quantification
3
Synthesis and Characterization of PFPMO
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PFPMO was synthesized according to a published procedure42 (link). Ultrapure agarose, CS, ALG, PEI (Mr = 55 000), tyrosinase (from mushroom, 4 276 units/mg), and dopamine were purchased from Sigma-Aldrich. Sodium citrate, silver nitrate, phenol, m-cresol, p-cresol, catechol and BPA were purchased from Beijing Chemical Reagent Co. Ltd.
4
Comet Assay for DNA Damage Evaluation
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This technique permits the detection and an evaluation of single-stranded DNA breaks. Eukaryotic cells are embedded in agarose gel on a microscopic slide, lysed by detergents and high salt at pH 10, and then electrophoresed for damage display which shows increased migration of the DNA from the nucleus towards the anode. Low-melting temperature agarose and ultra pure a garose, Triton X-100, sodium sarcosinate, ethylenediamine-tetra acetic acid disodium salt (Na-EDTA), Trizma base and ethidium bromide were obtained from Sigma Chemical Co. (St. Louis, MO, USA). Phosphate buffered saline without calcium and magnesium, RBMI 1640 medium (Gibco-BRL, Gaithersburg, MD, USA), Ficoll separating solution and trypan-blue were used in comet assay. Examination was done with a fluorescent microscope (BX 41, Olympus, Tokyo, Japan) equipped with an excitation filter of 510 nm and barrier filter of 590 nm. The migration was evaluated by observing and measuring the nuclear DNA, and 500 spots of DNA were examined and classified into three types: 1) normal spots with round shape; 2) damaged spots in which the length of the migrated fragments was less than or equal to the diameter of the basal nuclear DNA; and 3) strongly damaged spots where the length of the comet was greater than the diameter of the basal nuclear DNA [19 (link)].
5
Extraction and Analysis of Nucleic Acids
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Phenol, chloroform, RNase A, proteinase K, ultrapure agarose, and other general chemicals were purchased from Sigma-Aldrich (Prague, Czech Republic). Deoxynucleotides for polymerase chain reaction (PCR) and molecular weight standard for electrophoresis (ΦX174DNA/HaeIII digest) were products of New England Biolabs, Inc (Ipswich, MA). Taq-Purple DNA polymerase and Combi PPP Master Mix for PCR were supplied by Top-Bio s r.o. (Prague). Protein standards for immunoblotting were kindly provided by Prof Paul F. Hollenberg, University of Michigan, Ann Arbor, MI (P450 2B6) and Prof F. Peter Guengerich, Vanderbilt University, Nashville, TN (P450 3A4).
6
DNA Fragmentation Validation by Agarose Gel
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The fragmentation status of DNA (by UV or sonication) was validated with a 3% concentration of agarose gel electrophoresis. In total, 3 g of ultra-pure agarose (Sigma Aldrich) was dissolved in 100 mL of 1× Tris acetate-EDDTA (TAE) buffer. The mixture was gently brought to a boil until the solution had turned clear and the agarose had fully dissolved. Once cooled to 50–60 °C, 10 μL of SYBRSafe was added to the solution and mixed. The mixture was then poured into casting plate and left for 20 min until the gel fully polymerized. Once set, the gel was placed into the gel rig and submerged with the addition of 1× TAE. Prior to loading onto the gel, each DNA sample was mixed with 4× Loading buffer (40% w/v sucrose, 0.25% w/v bromophenol blue). The comb was removed and the samples were then added into the preset lanes. After sample loading, 6 μL of GeneRuler 100 bp DNA Ladder Plus was added for relative band size comparison. The gel was then resolved for ~90 min at 100 V before being visualized using a G-BOX gel documentation and analysis system.
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