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High-density oligonucleotide (oligo) arrays are a powerful tool for transcript profiling. Arrays based on GeneChip ®  technology are amongst the most widely used, although GeneChip ®  arrays are currently available for only a small number of plant and animal species. Thus, we have developed a method to improve the sensitivity of high-density oligonucleotide arrays when applied to heterologous species and tested the method by analysing the transcriptome of  Brassica oleracea  L., a species for which no GeneChip ®  array is available, using a GeneChip ®  array designed for  Arabidopsis thaliana  (L.) Heynh. Genomic DNA from  B. oleracea  was labelled and hybridised to the ATH1-121501 GeneChip ®  array.  Arabidopsis thaliana  probe-pairs that hybridised to the  B. oleracea  genomic DNA on the basis of the perfect-match (PM) probe signal were then selected for subsequent  B. oleracea  transcriptome analysis using a .cel file parser script to generate probe mask files. The transcriptional response of  B. oleracea  to a mineral nutrient (phosphorus; P) stress was quantified using probe mask files generated for a wide range of gDNA hybridisation intensity thresholds. An example probe mask file generated with a gDNA hybridisation intensity threshold of 400 removed > 68 % of the available PM probes from the analysis but retained >96 % of available  A. thaliana  probe-sets. Ninety-nine of these genes were then identified as significantly regulated under P stress in  B. oleracea , including the homologues of P stress responsive genes in  A. thaliana . Increasing the gDNA hybridisation intensity thresholds up to 500 for probe-selection increased the sensitivity of the GeneChip ®  array to detect regulation of gene expression in  B. oleracea  under P stress by up to 13-fold. Our open-source software to create probe mask files is freely available    and may be used to facilitate transcriptomic analyses of a wide range of plant and animal species in the absence of custom arrays.

Using genomic DNA-based probe-selection to improve the sensitivity of high-density oligonucleotide arrays when applied to heterologous species