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S. aureus DNA gyrase - for 100 assays

BHE16300198

This product includes 43 ul of 7.5 uM S. aureus gyrase.

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Reference 1. Asha M.K. et al, In vitro anti-Helicobacter pylori activity of a flavonoid rich extract of Glycyrrhiza glabra and its probable mechanisms of action, Journal of Ethnopharmacology, Vol.145(2), pp.581- 586 (2013).
2. Mora-Pale M et al, Antimicrobial mechanism of resveratrol-trans-dihydrodimer produced from peroxidase-catalyzed oxidation of resveratrol. Biotechnol Bioeng.Vol 112, pp2417–2428 (2015). 
Product Manual Datasheet
Protocols Description Assay Protocol using the S. aureus DNA Topoisomerase II (Gyrase) Assay Kit
The following assay protocol is based on a 96-well assay plate format using a standard black 96-well plate (Greiner 655076). For 384-well plate assays using a standard black 384-well plate (Matrix 4318), please reduce the reagent volumes proportionally.
 
1. Reaction:
The total volume of each reaction mixture is 40µl including: 19.6µl of H2O, 4µl of 10 x buffer, 8µl of 2 M potassium glutamate, 4µl of 10 x relaxed DNA (250µg/ml), 0.4µl of 7.5µM S. aureus gyrase, 4µl of 10 mM ATP. Incubate the reaction mixture at 37°C for 60 min.
 
Note: The final concentrations are 20 mM Tris-HCl, pH 8, 35 mM NH4OAc, 4.6 % glycerol, 1 mM DTT, 0.005% Brij35, 8 mM MgCl2, 400 mM potassium glutamate, 25µg/ml relaxed plasmid DNA, 1 mM ATP and 75 nM topoisomerase II. A negative control reaction can be the reaction mixture without addition of ATP. Magnesium is essential for the reaction and assay. EDTA should be avoided.
 
2. Assay
(1) Make 1 x H19 Dilution Buffer by dilution of the 10 x H19 Dilution Buffer 10-fold with water. Prepare the H19 dye by dilution of 1µl the 1500 x H19 dye stock solution with 1.5 ml of 1 x H19 Dilution Buffer (1500 x dilution). Mix the solution evenly by inverting the tube a few times.
(2) Mix 250µl of the freshly prepared H19 dye with each reaction solution (40µl). Incubate the mixture at room temperature for 5 min.
(3) Measure the fluorescence intensity at 535 nm using the excitation wavelength at 485 nm.
SKU Supplier No. Size Your price Quantity
BHE16300198 TOP2-100SA 100 assays $421.56
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