(A) Detecting allelic variation with high-density arrays. For nonduplicated regions of the genome, a minimum of 20 25-base oligonucleotide probes was chosen from yeast genomic sequence (S288c) for every annotated ORF (open reading frame = putative gene) in the yeast genome. Probes (only from predicted coding regions) were generally arranged on the array in order of their chromosome position. In addition to probes designed to be perfectly complementary to regions of yeast coding sequence (PM), probes containing a single base mismatch in the central position of the oligonucleotide were also synthesized in a physically adjacent position. The mismatch probes serve as background and nonspecific hybridization controls in other analyses. If probes complementary to YJM789 DNA fragments containing polymorphisms (*) are found on the array, decreases in signal intensity at these probes relative to the S96 signal may be observed when YJM789 DNA is hybridized to the array. The amount of signal decrease will depend on several factors, such as initial probe intensity and whether the probed fragment is completely absent in YJM789 or contains a small substitution. The location of the polymorphism within the probe sequence will also affect the observed intensity decrease. (B) Comparative genomic DNA hybridization patterns. Genomic DNA from two strains of S. cerevisiae, YJM789 and S96, was fluorescently labeled and hybridized to two different arrays. Scanned images of the arrays were collected, digitally colored red or green, and then electronically superimposed. A portion of the composite image is shown. Probes that hybridized to S96 DNA more efficiently than YJM789 DNA are red, and probes that hybridize to both DNA samples with equal intensity are yellow. A region that is completely deleted in YJM789 is indicated by an arrow. The figure closeup shows a region in which one of the mismatch features is bright green. Shotgun sequencing of YJM789 demonstrated that the actual sequence of YJM789 was complementary to the sequence of the oligonucleotide in the mismatch row and not to that in the perfect match row.
From: Winzeler EA, Richards DR, Conway AR, Goldstein AL, Kalman S, McCullough MJ, McCusker JH, Stevens DA, Wodicka L, Lockhart DJ, and Davis RW (1998) Direct allelic variation scanning of the yeast genome. Science 281: 1194-1197.