Hybridization of synthetic oligodeoxyribonucleotides to Φ_<i>X</i>174 DNA: the effect of single base pair mismatch

TLDR

Oligonucleotides are commonly used as probes to isolate specific cloned DNA sequences. The authors synthesized 11‑, 14‑, and 17‑base phosphotriester oligonucleotides complementary to the am‑3 mutation region of ΦX174 DNA and measured the thermal stability of the resulting duplexes. Hybridization of the oligonucleotides to am‑3 DNA produced duplexes with a single base‑pair mismatch that dissociate about 10 °C lower than the matched duplexes, enabling selective elimination of mismatched hybrids by adjusting the hybridization temperature.

Abstract

Oligodeoxyribonucleotides complementary to the DNA of the wild type (wt) bacteriophage Φ X 174 have been synthesized by the phosphotriester method. The ollgomers, 11, 14, and 17 bases long, are complementary to the region of the DNA which accounts for the am-3 point mutation. When hybridized to am-3 DNA, the oligonucleotides form duplexes with a single base pair mismatch. The thermal stability of the duplexes formed between wt and am-3 DNAs has been measured. The am-3 DNA: ollgomer duplexes dissociate at a temperature about 10°C lower than the corresponding wt DNA: ollgomer duplexes. This dramatic decrease in thermal stability due to a single mismatch makes it possible to eliminate the formation of the mismatched duplexes by the appropriate choice of hybridization temperature. These results are discussed with respect to the use of oligonucleotides as probes for the isolation of specific cloned DNA sequences.