Synthesis of modified nucleoside triphosphates and polymerase construction of functionalized nucleic acids
Novel efficient and expeditious approach for the construction of functionalized nucleic acids is being developed. It is based on aqueous-phase cross-coupling reactions of halogenated nucleoside triphosphates followed by enzymatic incorporation of the modified dNTPs by DNA polymerase. Primer extension experiments are used for the incorporation of the just one or several modified nucleotides in specific positions and, in combination with magnetoseparation of biotinylated template, to isolate single strand ONs. PCR is used to get larger DNA duplexes with a high degree of modifications. Terminal transferase is used for tail-labeling of 3'-end of ONs. Specific methods for introduction of one or several specific modifications into longer DNA and for production of larger amounts of modified ONs and DNA are under development.