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Cross-hybridization of repetitive sequences in genomic and expression arrays is reported to be suppressed with repeat-blocking nucleic acids (Cot-1 DNA). Contrary to expectation, we demon- strated that Cot-1 also enhanced non-specific hybrid- ization between probes and genomic targets. When added to target DNA, Cot-1 enhanced hybridization (2.2- to 3-fold) to genomic probes containing con- served repetitive elements. In addition to repetitive sequences, Cot-1 was found to be enriched for linked single copy (sc) sequences. Adventitious association between these sequences and probes distort quant- itative measurements of the probes hybridized to desired genomic targets. Quantitative microarray hybridization studies using Cot-1 DNA are also sus- ceptible to these effects, especially for probes that map to genomic regions containing conserved repet- itive sequences. Hybridization measurements with such probes are less reproducible in the presence of Cot-1 than for probes derived from sc regions or regions containing divergent repeat elements, a find- ing with significant ramifications for genomic and expression microarray studies. We mitigated the requirement for Cot-1 either by hybridizing with com- putationally defined sc probes lacking repeats or by substituting synthetic repetitive elements com- plementary to sequences in genomic probes.

Distortion of quantitative genomic and expression hybridization by Cot-1 DNA: mitigation of this effect