Covalent cross-linking of duplex DNA using 4-thio-2'-deoxyuridine as a readily modifiable platform for introduction of reactive functionality into oligonucleotides.
Full details of the template-directed covalent cross-linking of duplex oligodeoxynucleotides are presented. 4-Thio-2'-deoxyuridine was incorporated synthetically into a 17mer oligodeoxynucleotide, and the thiocarbonyl group of the modified base was alkylated with a variety of alpha-bromoacetyl-derivatized diamines. Covalent cross-linking was initiated by annealing the electrophilic probe oligomers with their complementary sequences, where a dG base was targeted at the position complementary to the modified 4-thio-2'-deoxyuridine. The sequence selectivity of cross-link formation as a function of tether topology and rigidity was examined, and the thermal stability of the modified duplexes was measured by UV melting experiments.
Coleman
R S
RS
Department of Chemistry, The Ohio State University, 100 West 18th Avenue, Columbus, OH 43210-1185, USA. coleman.184@osu.edu
Pires
R M
RM
D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D003432:Cross-Linking Reagents / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D004261:DNA Replication; D007700:Kinetics; D008956:Models, Chemical; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000378:metabolism*; D013891:Thiouridine / Q000031:analogs & derivatives* / Q000378:metabolism
D003432:Cross-Linking Reagents; D009841:Oligonucleotides; D013891:Thiouridine; C004274:4-thio-2'-deoxyuridine; D004247:DNA