Chemically Synthesized Nonradioactive DNA Probes. Final Project Report of Phase 1

摘要

The aim of the Phase I research project was to develop efficient chemical ways to introduce biotin into synthetic DNA fragments. Researchers investigated the solid-phase phosphoramidite one-stage method which allows biotinylation of DNA to be effected directly on the solid support. They succeeded in synthesizing two key compounds, N-monomethoxytrityl-6-amino-1-hexanol (compound 10) and N-monomethoxytrityl-o-(2-cyanoethoxy)diisopropylaminophosphinyl-6-amino- 1-hexanol (compound 11). These two compounds were characterized by NMR spectroscopy. By using compound 11, the researchers were able to synthesize and isolate a 21-mer (nt 1253) with hexylamine-linker at the 5' end position of the oligonucleotide chain, complementary to the P region of HBV (type AYW) genome. Biotinylation of the amine-linker oligonucleotide with N-biotinyl-w-aminocaproic ester was accomplished in a reasonable yield. The investigators further demonstrated the selectivity of the synthetic biotinylated oligonucleotide probe to localize HBV-DNA in HBV-producing cells in culture.