dATP


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Related to dATP: Dato, Ribonucleotide reductase

dATP

 (dē′ā′tē′pē′)
n.
One of the two purine nucleotides that are used to synthesize DNA.

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Table-1: Mean and standard deviation values of the clinical characteristics in control, high risk (HR) and preeclampsia group (PET) Characteristics Control Group (n=40) Age (years) 31.20 [+ or -] 5.84 BMI (kg/[m.sup.2]) 29.94 [+ or -] 6.05 Gestational Age (weeks) 31.17 [+ or -] 5.33 Haematocrit (%) 34.75 [+ or -] 4.30 Platelet Count 266.17 [+ or -] 84.83 ([10.sup.3]/[micro]l) sATP (mmHg) 113.56 [+ or -] 13.93 dATP (mmHg) 67.66 [+ or -] 9.38 S.
However, each of these reaction mixtures contained 20 nM of the template, 10 nM of each primer 5' -TATTGATTGTGAATTA(C/G)-3', and 100 [micro]M of a single dNTP (dCTP, dGTP, dATP, or dTTP).
Because this is done in real-time where the x-axis of the graph is time, the first 3 nucleotides dispensed (deoxythymidine triphosphate [dTTP], deoxyguanosine triphosphate [dGTP], and dCTP) do not elongate the growing strand because the first complementary nucleotide is dATP. Thus, there are no peaks on the pyrogram for these dispensed dNTPs (abbreviated T, G, and C in the graph) because they could not be incorporated into the growing strand.
For this, they used genetic engineering to create a strain of mice whose cells produced higher-than-normal levels of an enzyme called Ribonucleotide Reductase that converts the precursor of ATP, adenosine-5'-diphosphate or ADP, to dADP, which, in turn, is rapidly converted to dATP.
PCR requires DNA template, target specific primers, four dNTPs (dATP, dTTP, dCTP, and dGTP), DNA polymerase, magnesium, and buffer.
For SSR analysis, a reaction volume of 20 [micro]l was set up containing 2.0 ul of 10 x PCR buffer [50 mM of Tris (pH 8.3), 500 mM of KCl]; 1.5 mM of Mg[Cl.sub.2]; 0.2 mM each of dATP, dCTP, dGTP and dTTP (Fermentas, USA); 0.3 mM each of reverse and forward primer (GeneLink, USA); one units of Taq DNA polymerase (Fermentas, USA); and 25 ng of genomic DNA as a template.
The supernatant was transferred to a fresh tube, the cDNA reamplified using the same primers and similar PCR conditions except radio-labeled dATP was not used and the dNTP concentrations were increased to 250 [micro]M.
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For labeling with Cy3-dATP, 0.5 mM of each dCTP, dGTP, and dTTP and 0.08 mM dATP (Qiagen, Hilden, Germany) were used instead of 0.5 mM dTTP.
A total of 50 of the first reaction mixture contained (1x) PCR buffer with 1.5 mM Mg[Cl.sub.2] (Invitrogen, USA), 200 [micro]M of dNTP (Invitrogen, USA) containing each of dATP, dCTP, dGTP, dTTP the PCR primers, 2.5 U of Taq DNA Polymerase (Intron, Korea), and 1 [micro]l of DNA template.
PCR amplification was carried out with PCR primers (20 pmol/L each per 50 [micro]L reaction) (Integrated DNA Technologies, Coralville, IA, USA) (Table) and 1 [micro]L genomic DNA (extracted from overnight Luria-Bertani broth cultures according to PureGene DNA isolation kit instructions [Gentra Systems, Minneapolis, MN, USA] and dissolved in 50 [micro]L 10 mmol/L Tris, pH 8.3) in a PCR cocktail containing 1x PCR buffer, 1.5 mmol/L Mg[Cl.sub.2], 1 U Vent exo(-) polymerase from New England BioLabs (Beverly, MA, USA), and 200 [micro]mol/L each dATP, dGTP, dTTP, and dCTP.
For each run, a master mix was prepared with 1 x SYBR Green buffer containing 5 mM Mg[Cl.sub.2]; 200 mM dATP, dCTP, and dGTP; 400 mM dUTP; 1.25 U AmpliTaq Gold DNA polymerase (Applied Biosystems, Courtaboeuf, France); and 300 nM of each primer: EXIIc sense primer, 5' TGA GGT CAA GGA ACA CAA GA 3', exon II specific (positions 9-28); and EXIII antisense primer, 5' ATC CAC AGG AAT CTG CCG TG 3', for exon III (positions 211-230) (Corbin et al.