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The enzymatic incorporation of deoxyribonucleoside triphosphates by a thermostable, 3′→5′ exonuclease deficient mutant of the Tgo DNA polymerase was studied for PCR-based high-density labeling of 217-bp ‘natural’ DNA in which fluorescent-dUTP was substituted completely for the normal dTTP. The amplified DNA carried two different sorts of tethered dye molecules. The rhodamine-green was used for internal...
We describe here the enzyme-catalyzed, low-density labeling of DNAs with fluorescent dyes. Firstly, for ‘natural’ template DNAs, dNTPs were partially substituted in the labeling reactions by the respective fluorophore-bearing analogs. The DNAs were labeled by PCR using Taq DNA polymerase. The covalent incorporation of dye-dNTPs decreased in the following order: rhodamine-green-5-dUTP (Molecular Probes,...
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