The plasmid DNA damage properties of isolated compounds were analyzed employing the agarose gel electrophoresis according to our previous studies [8 (link)]. The results were monitored by a BioRad Gel Doc XR system and analyzed using Image Lab Version 4.0.1 Software Program. Briefly, the reaction mixture [isolated compounds in Tris–HCl buffer (50 mM, pH 7) containing dimethyl sulfoxide (final concentration 1%), supercoiled pBR322 plasmid DNA (Thermo Scientific, SD0041)] were preincubated for 30 min at 37 °C. Afterwards, loading buffer (bromophenol blue) (0.2%) (Sigma, B0126), xylene cyanol (0.2%) (Sigma, X4126), glycerol (30%) (Sigma, G5516), and sodium dodecyl sulfate (4.5%) (Sigma, L3771) were added to the samples. The samples were loaded on 0.8% (w/v) agarose (Sigma, A9539) gel with ethidium bromide (Sigma, E7637) staining in Tris–acetic acid–EDTA (TAE) (1X) buffer. Electrophoresis was performed at 100 V for 90 min using Wide Mini ReadySub-Cell GT Horizontal Electrophoresis System (BioRad).
B0126
Manufactured by Merck Group
B0126 is a laboratory instrument designed for general-purpose use in scientific research and analysis. It is a high-performance device capable of performing a range of tasks within a controlled laboratory environment. The core function of B0126 is to enable precise measurements and data collection, supporting the research and development activities of scientists and technicians.
Automatically generated - may contain errors
Lab products found in correlation
G5516, by Merck Group (2 mentions)
A9539, by Merck Group (1 mentions)
L3771, by Merck Group (1 mentions)
E7637, by Merck Group (1 mentions)
Pbr322 plasmid dna, by Thermo Fisher Scientific (1 mentions)
Gel doc xr system, by Bio-Rad (1 mentions)
Tween 20, by Merck Group (1 mentions)
Clarity western ecl substrate, by Bio-Rad (1 mentions)
Trans blot turbo system, by Bio-Rad (1 mentions)
D9779, by Merck Group (1 mentions)
Las 1000plus, by Fujifilm (1 mentions)
Pepstatin a, by Roche (1 mentions)
2 protocols using b0126
1
Plasmid DNA Damage Analysis by Gel Electrophoresis
2
Western Blot Protein Detection
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Cells were lysed in Laemmli sample buffer (125 mM Tris, pH 6.7, 20% glycerol, 140 mM SDS [Sigma-Aldrich, G5516 and L3771]) supplemented with 3x complete protease inhibitor cocktail (Roche, 11697498001) and 40 µg/ml pepstatin A (Roche, 10359053001) at 10000 cells/µl. After addition of 0.1 M dithiothreitol and bromophenol blue (Sigma-Aldrich, D9779 and B0126), boiling and separation by 4–20% gradient SDS-PAGE, proteins were transferred onto polyvinylidene difluoride (PVDF) membranes using Bio-Rad Trans-Blot Turbo system. Membranes were blocked with DPBS containing 5% milk and 0.1% Tween-20 (Sigma-Aldrich, 274348) and labeled with the antibodies listed above. The signal was detected with Clarity Western ECL Substrate (Bio-Rad, 170–5061), and Luminescent Image Reader (LAS-1000Plus, Fujifilm, Tokyo, Japan), and quantified by densitometry with ImageJ software.
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