Accelerated hybridization of oligonucleotides to duplex DNA.
We report two strategies for accelerating the hybridization of oligonucleotides to DNA. We demonstrate that oligodeoxyribonucleotides and peptide nucleic acid oligomers hybridize to inverted repeats within duplex DNA by D-loop formation. Oligonucleotides and duplex template form an active complex, which can be recognized by T7 DNA polymerase to prime polymerization. Quantitation of polymerization products allowed the rate of hybridization to be estimated, and peptide nucleic acid oligomers and oligonucleotide-protein adducts anneal with association constants 500- and 12,000-fold greater, respectively, than the analogous unmodified oligonucleotides. Together, these results indicate that sequences within duplex DNA can be targeted by Watson-Crick base pairing and that chemical modifications can dramatically enhance the rate of strand association. These findings should facilitate targeting of oligomers for priming DNA polymerization, the detection of diagnostic sequences, and the disruption of gene expression. The observed acceleration of hybridization may offer a new perspective on the ability of RecA or other proteins to accelerate strand invasion.
Iyer
M
M
Howard Hughes Medical Institute, Department of Pharmacology, University of Texas Southwestern Medical Center at Dallas 75235, USA.
Norton
J C
JC
Corey
D R
DR
D001483:Base Sequence; D004247:DNA / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D009841:Oligonucleotides / Q000235:genetics*; D011506:Proteins / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid
D009841:Oligonucleotides; D011506:Proteins; D004247:DNA