Characterization of fluorescent nucleoside triphosphates by capillary electrophoresis with laser-induced fluorescence detection: action of alkaline phosphatase and DNA polymerase.
A method of analysis of fluor-labeled nucleoside triphosphates based on alkaline phosphatase-catalyzed sequential cleavage of phosphate groups with monitoring of all fluorescent species by capillary electrophoresis with laser-induced fluorescence detection is presented. The method allows determination of the purity of the triphosphate samples as well as the relative amounts of the lower phosphate contaminants. The ability of one of the fluor-labeled nucleoside triphosphates to serve as polymerase substrate was verified by labeling DNA restriction fragments by the method of filling recessed 3'-ends using DNA polymerase Klenow fragment.
Evangelista
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RA
Advanced Technology Center, Beckman Instruments Inc., Fullerton, California 92634, USA.
Liu
M S
MS
Rampal
S
S
Chen
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FT
D000255:Adenosine Triphosphate / Q000032:analysis*; D000469:Alkaline Phosphatase / Q000737:chemistry*; D003570:Cytidine Triphosphate / Q000032:analysis*; D004274:DNA, Recombinant / Q000737:chemistry; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry*; D007834:Lasers; D013050:Spectrometry, Fluorescence; D014544:Uridine Triphosphate / Q000032:analysis*
D004274:DNA, Recombinant; D003570:Cytidine Triphosphate; D000255:Adenosine Triphosphate; D004259:DNA-Directed DNA Polymerase; D000469:Alkaline Phosphatase; D014544:Uridine Triphosphate