Successful genetic mapping is dependent upon a high-density set of markers. Therefore, tools for high-throughput discovery of genetic variation are essential. The most abundant genetic marker is the single-nucleotide polymorphism (SNP). However, except for model organisms, genomic information is still limited. Although high-throughput genomic sequencing technologies are becoming relatively inexpensive, only low-throughput genetic markers are accessible (e.g., simple sequence repeats). The use of sequencing for the discovery and screening of high-density genetic variation in whole populations is still expensive. Alternatively, hybridization of genomic DNA (gDNA) on a reference (either genome or transcriptome) is an efficient approach for genetic screening without knowing the alleles in advance (Borevitz et al. Proc Natl Acad Sci USA 104:12057-12062). We describe a protocol for the design of probes for a high-throughput genetic-marker discovery microarray, termed single feature polymorphism (SFP) array. Starting with consensus cDNA sequences (UniGenes), we use OligoWiz to design T (m)-optimized 50-bp long oligonucleotide probes (Ophir et al. BMC Genomics 11:269, 2010). This design is similar to expression arrays and we point out the differences.