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Biotechniques ; 64(1): 21-23, 2018 01 01.
Article in English | MEDLINE | ID: mdl-29384073

ABSTRACT

Previously, we reported a method for facile purification of oligonucleotides labeled with hydrophobic dyes, based on the solubility difference between the hydrophilic DNA and unreacted dye. Here, we present a new purification method applicable to any dye regardless of its hydrophobicity. We exploited the population shift of a fluorescent dye in a low-pH aqueous solution from its anionic form toward its neutral form. When the pH of an aqueous solution containing dye-labeled DNA and unreacted free dye is lowered, and the solution is mixed with a hydrophobic organic solvent (butanol), the neutral free dye is preferentially dissolved in the organic phase, leaving behind the hydrophilic dye-labeled DNA in the aqueous phase. We experimentally verified that our new method results in high yields of dye-labeled oligonucleotides and the efficient removal of free dye.


Subject(s)
Carbocyanines/isolation & purification , DNA/isolation & purification , Fluorescent Dyes/isolation & purification , Oligonucleotides/isolation & purification , Chemical Precipitation , Chromatography, High Pressure Liquid , Hydrogen-Ion Concentration , Hydrophobic and Hydrophilic Interactions , Phase Transition , Solubility , Solutions , Water/chemistry
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