Tutorial section: Designing microarray oligonucleotide probes

INTRODUCTION DNA microarrays are one of a few technologies that enable the monitoring of thousands of gene expression levels in parallel. In general, microarrays comprise a solid planar substrate on which an ordered array of probes has been deposited. Each probe represents a gene or transcript of interest and there are essentially three main types of probe: polymerase chain reaction (PCR) products of cDNA clones, ‘long’ oligonucleotides and ‘short’ oligonucleotides. Synthetic oligonucleotides, long or short, offer a number of practical advantages over cDNA clones for constructing arrays, making oligonucleotides an attractive alternative. The ease of automated synthesis, in comparison to the preparation of libraries of cDNA molecules, is a major attraction. In addition, synthetic probes offer the possibility of precise control over the composition and size of each probe. There is also the possibility of using nonnatural nucleic acid analogues in the construction of oligonucleotide arrays, which may have advantages. Peptide nucleic acids (PNA), for example, have a neutral backbone that overcomes the usual, mutual repulsion of a duplex of natural phosphodiesters. Hence PNA has a higher binding affinity for DNA and shows greater stringency in hybridisation than DNA. Finally, oligonucleotide probes can be designed based on published sequences in a wide variety of databases and, consequently, there has been a substantial interest in the development of bioinformatics tools that allow for the design of oligonucleotide probes from these databases.