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Journal of Virology, July 1999, p. 5757-5766, Vol. 73, No. 7
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

DNA Microarrays of the Complex Human Cytomegalovirus Genome: Profiling Kinetic Class with Drug Sensitivity of Viral Gene Expressiondagger

James Chambers,1 Ana Angulo,2 Dhammika Amaratunga,1 Hongqing Guo,1 Ying Jiang,1 Jackson S. Wan,1 Anton Bittner,1 Klaus Frueh,1 Michael R. Jackson,1 Per A. Peterson,1 Mark G. Erlander,1 and Peter Ghazal2,*

Departments of Immunology and Molecular Biology, Division of Virology, The Scripps Research Institute, La Jolla, California 92037,2 and The R. W. Johnson Pharmaceutical Research Institute, San Diego, California 921211

Received 28 December 1998/Accepted 9 April 1999

We describe, for the first time, the generation of a viral DNA chip for simultaneous expression measurements of nearly all known open reading frames (ORFs) in the largest member of the herpesvirus family, human cytomegalovirus (HCMV). In this study, an HCMV chip was fabricated and used to characterize the temporal class of viral gene expression. The viral chip is composed of microarrays of viral DNA prepared by robotic deposition of oligonucleotides on glass for ORFs in the HCMV genome. Viral gene expression was monitored by hybridization to the oligonucleotide microarrays with fluorescently labelled cDNAs prepared from mock-infected or infected human foreskin fibroblast cells. By using cycloheximide and ganciclovir to block de novo viral protein synthesis and viral DNA replication, respectively, the kinetic classes of array elements were classified. The expression profiles of known ORFs and many previously uncharacterized ORFs provided a temporal map of immediate-early (alpha ), early (beta ), early-late (gamma 1), and late (gamma 2) genes in the entire genome of HCMV. Sequence compositional analysis of the 5' noncoding DNA sequences of the temporal classes, performed by using algorithms that automatically search for defined and recurring motifs in unaligned sequences, indicated the presence of potential regulatory motifs for beta , gamma 1, and gamma 2 genes. In summary, these fabricated microarrays of viral DNA allow rapid and parallel analysis of gene expression at the whole viral genome level. The viral chip approach coupled with global biochemical and genetic strategies should greatly speed the functional analysis of established as well as newly discovered large viral genomes.

* Corresponding author. Mailing address: Departments of Immunology and Molecular Biology, Division of Virology, The Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, CA 92037. Phone: (619) 784-8678. Fax: (619) 784-9272. E-mail: ghazal{at}scripps.edu.

dagger This is publication no. 12111-IMM from The Scripps Research Institute.

Journal of Virology, July 1999, p. 5757-5766, Vol. 73, No. 7
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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