Oligonucleotide synthesis on maleic anhydride copolymers covalently bound to silica spherical support and characterization of the obtained conjugates

摘要

A new route was proposed to make polymer-oligonucleotide conjugates of potential applications in diagnostics. It consisted in direct synthesis of oligonucleotides onto controlled pore glass surface grafted with poly(maleic anhydride-alt-methyl vinyl ether) (P[MAMVE]) or poly(maleic anhydride-alt-ethylene) (P[MAE]). The anhydride moieties were used for both the covalent coupling of the copolymer via ester bond and binding of 5'-dimethoxytrityl thymidine 3'-(6-aminohexyl phosphate) (I) initiator of oligodeoxynucleotide (ODN) synthesis via amide bond. The difference of stability between ester and amide links under basic treatment was used for the selective cleavage of (polymer-oligonucleotide) conjugates after DNA synthesis completion. We succeeded in grafting functionalized copolymer onto silica surface and Polythymidine 26-mer ODN was performed. After concentrated ammonium hydroxide treatment, conjugate crude materials were characterized by size exclusion chromatography coupled to multiangle laser light scattering detection. The number average molecular weight (Mn) for conjugate with P[MAMVE] was abnormally lower than expected and was assigned to polymer degradation using high pH conditions. Such a phenomenon did not occur with P[MAE]-polythymidine conjugate. However, in both cases, parasite ODN synthesis was also evidenced, which was attributed to thymidine phosphoramidite adsorption side reaction during DNA synthesis. (C) 1998 John Wiley & Sons, Inc. [References: 18]