Chemical and enzymatic biotin-labeling of oligodeoxyribonucleotides

Abstract

Biotin has been converted to 2-(biotinylamido)ethanol and condensed to phosphorylated oligonucleotides in a solid phase synthesis. The 5'-biotinylated oligonucleotides were enzymatically coupled to other DNA fragments by T4 DNA ligase or T4 RNA ligase. The hybridization properties of such biotin-labeled oligonucleotide probes were studied.

Evaluation of the 2 NH2A—T Pair in Hybridization, I Synthesis of the DNA/RNA Hybrid Oligomers Containing 2-Aminoadenosines

“…In spite of the reported stabilization of the DNA duplex by the ribonucleotide substitution [11] and the predicted efficacy of hydrogen bonding using 2 NH 2 A-T pair, 2-aminoadenosine was not effective in our case of "hybrid probe". The "RNA probe" is more effective in hybridization than the "DNA probe", and recently in vitro RNA probe preparation method [21] has been developed.…”

“…It is well understood that the glycosidic bond of the purine ribonucleoside is more stable than that of the corre-sponding 2'-deoxynucleoside. Moreover, Kempe et al [11] have recently reported that DNA oligomers containing a ribonucleotide (DNA/RNA hybrid) cause the elevation of the Tm value to a certain extent in comparison to the corresponding DNA oligomers. Miles et al [12] reported that elevation of the Tm value by the 2-amino group is much smaller in the deoxy than in the ribo series.…”

New Approaches to the Detection of Microbial Plant Pathogens

“…Advances in nucleic acid chemistry have led to the development of various non-radioactive probes for nucleic acid hybridization (Kempe et al, 1985 Kincaid and Nightingale, 1988). The basic strategies include:…”

Strategies for introducing non-radioactive labels during the automated sequence analysis of nucleic acids

“…On the other hand, as far as an enzymatic addition of a label is concerned, the 5' OH is particularly inert. One illustrative approach was an attempted ligation of a phosphorylated deoxyoligonucleotide to an biotinylated trinucleotide terminating in ribo C (Kempe et al, 1985). In the presence of a total of 10µg RNA ligase and over a incubation period of 96 h, a 20 % yield of the desired biotinylated oligonucleotide was obtained.…”