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Home / Assay Kits

E. coli DNA Primase Assay Kit

SKU : BHE16300079

This product includes 600 ul of 10 x assay buffer, 45 ul of 100 x DNA template, 45 ul of 100 x NTP mix and 850 ul of 10 x fluorescence dye for 100 assays of E. coli DNA primase reactions in a 96-well plate format or 200 assays in 384-well assay format. The kit does not include the enzyme.

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Mfr.No.	EGA100K, EGA100KE
Description	The bacterial DNA primase (DnaG) synthesizes RNA primers at the DNA replication fork where the DNA helicase (DnaB) unwinds the double strand DNA. The Bacterial DNA Primase Assays are based on measurement of the RNA primers synthesized by the DNA primase in the presence of the DNA temperate and NTPS. DNA helicases dramatically stimulate the activities of gram-negative bacterial primases (e.g. E. coli and H. influenzae) but not gram-positive bacterial primases (S. aureus and S. pneumoniae). The assays can be performed in a 96-well plate or 384-well plate format for high throughput screening of DNA primase inhibitors. The E. coli DNA Primase Assay Kit (Catalog No. EGA100K) includes 600 µl of 10 x assay buffer, 45 µl of 100 x DNA template, 45 µl of 100 x NTP mix and 850 µl of 10 x fluorescence dye for 100 assays of E. coli DNA primase reactions in a 96-well plate format or 200 assays in 384-well assay format. The kit does not include the enzyme.
Drug target assays	Antibacterial drug targets

Reference	Lacriola CJ et al, Inhibition of DNA replication in Staphylococcus aureus by tegaserod. The Journal of Antibiotics. 70, 918–920 (2017).

Product Manual	Datasheet
Protocols Description	Assay Protocol 1.Reagent preparation: 10 x DNA: dilute the 100 x DNA with water. 10 x enzyme: Dilute the 100 x enzyme stock with the 1 x assay buffer to make the 10 x enzyme. 10 x NTP mix: dilute the 100 x NTP 10-fold with water. 1 x fluorescence dye: dilute the 10 x fluorescence dye 10-fold with water. 2. Reaction: The total volume of each reaction mixture is 40µl including 24µl of H2O, 4µl of 10 x Buffer, 4µl of 10 x DNA template, 4µl of 10 x enzyme, 4µl of 10 x NTP mix. Incubate the reaction mixture at 37°C for 60 min. Note: The assay solution is composed of 10 mM HEPES, pH 7.5, 5 mM magnesium acetate, 0.5 mM DTT, 0.003% Brij-35, 100 nM DNA, 0.5 mM NTPs, 100 nM enzyme. 3. Detection: Add 80µl of the 1 x fluorescence dye into the 40µl of the reaction mixture. Incubate for 5 min. Measure the fluorescence intensity at 535 nm using the excitation wavelength at 485 nm.

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Combination SKU	Supplier No.	Specifications	Your price	Quantity
BHE16300079-1	EGA100K	enzyme not included	$365.40		Add to Cart
BHE16300079-2	EGA100KE	enzyme included	$613.14		Add to Cart

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