Isostable DNA.
The high fidelity detection of multiple DNA sequences in multiplex assays calls for duplexes whose stability is independent of sequence (isostable DNA), forming under universally stringent conditions. Nature did not evolve DNA to form isostable duplexes. Here we report how probe strands can be modified so that an all-A/T target strand is bound with the same or slightly higher affinity than the corresponding all-G/C strand with the same sequence of purines and pyrimidines. We refer to these probes that feature covalently attached ligands as "decorated nucleic acids". Caps, intercalators, and locks were used to stabilize A/T duplexes, and N4-ethylcytosine residues were employed to tune down the stability of G/C duplexes without significantly affecting base pairing selectivity. Near-isostability was demonstrated in solution and on microarrays of high and low density. Further, it is shown that hybridization results involving decorated probes on microarrays can be predicted on the basis of thermodynamic data for duplex formation in solution. Predictable formation of isostable DNA not only benefits microarrays for gene expression analysis and genotyping, but may also improve the sequence-specificity of other applications that rely on the massively parallel formation of Watson-Crick duplexes.
Ahlborn
Carolin
C
Institut für Organische Chemie, Universität Karlsruhe (TH), Karlsruhe, Germany.
Siegmund
Karsten
K
Richert
Clemens
C
D000225:Adenine / Q000737:chemistry; D020029:Base Pairing; D004247:DNA / Q000737:chemistry*; D004355:Drug Stability; D005453:Fluorescence; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000737:chemistry; D013816:Thermodynamics; D013936:Thymidine / Q000737:chemistry
D009841:Oligonucleotides; D004247:DNA; D000225:Adenine; D013936:Thymidine