Bacterial artificial chromosome-emulation oligonucleotide arrays for targeted clinical array-comparative genomic hybridization analyses

Purpose: The goal of this work was to test the ability of oligonucleotide-based arrays to reproduce the results of focused bacterial artificial chromosome (BAC)-based arrays used clinically in comparative genomic hybridization experiments to detect constitutional copy number changes in genomic DNA. Methods: Custom oligonucleotide (oligo) arrays were designed using the Agilent Technologies platform to give high-resolution coverage of regions within the genome sequence coordinates of BAC/P1 artificial chromosome (PAC) clones that had already been validated for use in previous versions of clone arrays used in clinical practice. Standard array-comparative genomic hybridization experiments, including a simultaneous blind analysis of a set of clinical samples, were conducted on both array platforms to identify copy number differences between patient samples and normal reference controls. Results: Initial experiments successfully demonstrated the capacity of oligo arrays to emulate BAC data without the need for dye-reversal comparisons. Empirical data and computational analyses of oligo response and distribution from a pilot array were used to design an optimized array of 44,000 oligos (44K). This custom 44K oligo array consists of probes localized to the genomic positions of >1400 fluorescence in situ hybridization-verified BAC/PAC clones covering more than 140 regions implicated in genetic diseases, as well as all clinically relevant subtelomeric and pericentromeric regions. Conclusions: Our data demonstrate that oligo-based arrays offer a valid alternative for focused BAC arrays. Furthermore, they have significant advantages, including better design flexibility, avoidance of repetitive sequences, manufacturing processes amenable to good manufacturing practice standards in the future, increased robustness because of an enhanced dynamic range (signal to background), and increased resolution that allows for detection of smaller regions of change.