Applications of DNA tiling arrays to experimental genome annotation and regulatory pathway discovery Paul Bertone 1 , Mark Gerstein 2 & Michael Snyder 1,2 1 Department of Molecular, Cellular, and Developmental Biology Yale University, New Haven, CT 06520-8103, USA; Tel: +1(203) 432-5405; Fax +1(203) 432-5175; E-mail: paul.bertone@yale.edu; 2 Department of Molecular Biophysics and Biochemistry Yale University, New Haven, CT 06520-8114, USA Key words: chromatin immunoprecipitation, gene expression, microarrays, regulatory network, transcript mapping Abstract Microarrays have become a popular and important technology for surveying global patterns in gene expression and regulation. A number of innovative experiments have extended microarray applications beyond the measurement of mRNA expression levels, in order to uncover aspects of large-scale chromosome function and dynamics. This has been made possible due to the recent development of tiling arrays, where all non-repetitive DNA comprising a chromosome or locus is represented at various sequence resolutions. Since tiling arrays are designed to contain the entire DNA sequence without prior consultation of existing gene annotation, they enable the discovery of novel transcribed sequences and regulatory elements through the unbiased interrogation of genomic loci. The implementation of such methods for the global analysis of large eukaryotic genomes presents significant technical challenges. Nonetheless, tiling arrays are expected to become instrumental for the genome-wide identification and characterization of functional elements. Combined with computational methods to relate these data and map the complex interactions of transcriptional regulators, tiling array experiments can provide insight toward a more comprehensive under- standing of fundamental molecular and cellular processes. Introduction It is widely recognized that the availability of a complete genome sequence can significantly enhance our ability to analyse biological phe- nomena and elucidate molecular and cellular function. Beyond the initial determination of the DNA sequence, the most valuable resource pro- duced by genome mapping efforts entails a com- prehensive catalogue of functional elements that encompass the genetic repertoire of an organism. Methods for the global analysis of gene expres- sion include subtractive hybridisation (Hedrick et al. 1984), differential display (Liang & Pardee 1992), and representational difference analysis (Hubank & Schatz 1994). While these techniques are useful for characterizing differences in mRNA transcript populations, they are unable to generate comprehensive gene expression profiles. The genome-wide identi¢cation of transcribed sequences was made possible with the develop- ment of the SAGE (serial analysis of gene expres- sion) technique (Velculescu et al. 1995). SAGE enables the quantitative estimation of mRNA expression levels by sampling short (10^14mer) sequences of transcribed messages, and using these to deduce the identity of the speci¢c transcripts from which they are derived. The advantages of Chromosome Research 13: 259–274, 2005. 259 # 2005 Springer. Printed in the Netherlands