Identification of nucleotides with identical fluorescent labels based on fluorescence polarization in surfactant solutions.
A solution-phase steady-state polarization-based method for discriminating among the four DNA nucleotides labeled identically with tetramethylrhodamine is described and demonstrated. Labeled nucleotides were dissolved in buffered surfactant solutions. In room temperature 4.5 mM Triton X-100 solutions at neutral pH, the measured steady-state polarizations of tetramethylrhodamine-labeled dATP, dCTP, dGTP and dUTP were 0.261 +/- 0.003, 0.112 +/- 0.003, 0.288 +/- 0.003, and 0.147 +/- 0.003, respectively. A blind test of 40 samples showed no errors in classification based on polarization. The reproducibility obtained during this study demonstrates that the four dye-labeled nucleotides can be discriminated with more than 99.8% confidence.
Yan
Y
Y
Department of Chemistry and Biochemistry, University of South Carolina, Columbia 29208, USA.
Myrick
M L
ML
D003838:Deoxyadenine Nucleotides / Q000032:analysis / Q000145:classification; D003845:Deoxycytosine Nucleotides / Q000032:analysis / Q000145:classification; D003848:Deoxyguanine Nucleotides / Q000032:analysis / Q000145:classification; D003856:Deoxyuracil Nucleotides / Q000032:analysis / Q000145:classification; D005454:Fluorescence Polarization / Q000379:methods*; D005456:Fluorescent Dyes; D015394:Molecular Structure; D009711:Nucleotides / Q000032:analysis / Q000145:classification*; D017830:Octoxynol; D012235:Rhodamines; D012996:Solutions; D013501:Surface-Active Agents
D003838:Deoxyadenine Nucleotides; D003845:Deoxycytosine Nucleotides; D003848:Deoxyguanine Nucleotides; D003856:Deoxyuracil Nucleotides; D005456:Fluorescent Dyes; D009711:Nucleotides; D012235:Rhodamines; D012996:Solutions; D013501:Surface-Active Agents; C027078:deoxyuridine triphosphate; C024107:2'-deoxycytidine 5'-triphosphate; C005358:tetramethylrhodamine; C029603:deoxyguanosine triphosphate; D017830:Octoxynol; C026600:2'-deoxyadenosine triphosphate