Synthesis of full-length oligonucleotides: cleavage of apurinic molecules on a novel support.
The synthesis of oligodeoxynucleotides is marred by several problems that contribute to the formation of defective molecules. This in turn seriously limits the usefulness of such reagents in DNA diagnostics, molecular cloning, DNA structural analysis and in antisense therapy. In particular, depurination reactions during the cyclical steps of synthesis lead to strand scission during cleavage of the completed molecules from the support. Here we present a remedy to this problem: a novel disiloxyl linkage that connects oligonucleotides to the support withstands reaction conditions that allow the removal of the 5' parts of any depurinated molecules. This ensures that all molecules that preserve the 5' protecting group when cleaved from the support will have both correct 3'- and 5'-ends. We demonstrate the application of the support for synthesis of padlock probe molecules.
Kwiatkowski
M
M
Department of Medical Genetics, Uppsala University, Sweden. marek.kwiatkowski@medgen.uu.se
Nilsson
M
M
Landegren
U
U
D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D011088:DNA Ligases / Q000378:metabolism; D004355:Drug Stability; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D015394:Molecular Structure; D015345:Oligonucleotide Probes / Q000737:chemistry; D009841:Oligonucleotides / Q000138:chemical synthesis*; D011687:Purines / Q000737:chemistry*; D012833:Siloxanes / Q000138:chemical synthesis
D007202:Indicators and Reagents; D015345:Oligonucleotide Probes; D009841:Oligonucleotides; D011687:Purines; D012833:Siloxanes; C031036:1,3-dichlorotetraisopropyldisiloxane; D011088:DNA Ligases