Esterification of terminal phosphate groups in nucleic acids with sorbitol and its application to the isolation of terminal polynucleotide fragments.
The exposure of mono- and polynucleotides to 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide and high concentrations of sorbitol results in the esterification of their monosubstituted phosphate groups. The presence of the sorbitol moiety permits these derivatives to bind strongly at pH 8.7 to columns of chromatographic supports containing the dihydroxyboryl group and to be subsequently released by elution with buffers at pH 5.5. The procedure constitutes a method for the isolation of polynucleotide fragments arising from the terminals of nucleic acids. A new method for the preparation of the chromatographic supports involves the synthesis of the 1,3-propanediol cyclic ester of m-[[3-(N-succinimidoxycarbonyl)propanoyl]amino]benzeneboronic acid and its condensation with aminoethylcellulose or amino-ethylpolyacrylamide. The reagent is readily prepared by reaction of N-[m-(dihydroxyboryl)phenyl)]succinamic acid with 1,3-propanediol to protect the boronate moiety followed by esterification with N-hydroxysuccinimide in the presence of dicyclohexylcarbodiimide.
D000818:Animals; D002417:Cattle; D004247:DNA; D004952:Esters; D005022:Ethyldimethylaminopropyl Carbodiimide; D007202:Indicators and Reagents; D008722:Methods; D009838:Oligodeoxyribonucleotides / Q000302:isolation & purification*; D009841:Oligonucleotides / Q000302:isolation & purification*; D009843:Oligoribonucleotides / Q000302:isolation & purification*; D012343:RNA, Transfer; D012441:Saccharomyces cerevisiae; D013012:Sorbitol; D013950:Thymus Gland
D004952:Esters; D007202:Indicators and Reagents; D009838:Oligodeoxyribonucleotides; D009841:Oligonucleotides; D009843:Oligoribonucleotides; D013012:Sorbitol; D004247:DNA; D012343:RNA, Transfer; D005022:Ethyldimethylaminopropyl Carbodiimide