A Direct Glimpse of Cross-Hybridization: Background-Passified Microarrays That Allow Mass-Spectrometric Detection of Captured Oligonucleotides

摘要

The generation of DNA microarrays or DNA chips[1], [2] has made possible the massively parallel detection of DNA and RNA sequences. The ability to detect DNA or RNA sequence selectivity with DNA chips relies on the formation of Watson-Crick duplexes between immobilized strands (probes) and the target strands in solution. High-density DNA chips feature up to 250 000 spots and allow gene expression to be monitored on the level of entire genomes.[3], [4] Duplex formation between DNA strands with only partial complementarity (cross-hybridization) is difficult to suppress at this level of complexity. As conventional DNA chip experiments use the same fluorophore for labeling all sequences from one biological sample, cross-hybridization can lead to false positive results, threatening the reliability of chip experiments. This is particularly true when single-base resolution is required, either because single-nucleotide polymorphisms have to be detected or because the expression of closely related genes has to be monitored.[5] Single-mismatch resolution is also important for control sequences employed on some chips for background correction,[6] an approach that has been challenged.