|
 |
 |
|
|
|
 |
|
|
1. What are DNA microarrays?
A gene consists of double stranded DNA molecules that form a helical structure. The double stranded DNA molecules unzip into two complementary strands when heated. Upon cooling, each of these can recombine or bind to a new DNA molecule that contains the same sequence as the complementary strand. This process of two complementary strands binding together is called hybridization and this reaction is used to detect DNA or RNAs of interest (targets) on the surface of the DNA microarray. Certain oligonucleotides (short chains of nucleic acids) or cDNAs (complementary DNAs synthesized from mRNA) are attached or synthesized on the surface of the DNA microarray to bind their complementary strands. These oligonucleotides and cDNA are called probes, and the targeted DNAs or RNAs can be detected by hybridization reactions with probes on the surface of the DNA microarray. A DNA microarray contains tens of thousands of oligonucletides or cDNAs attached on the surface; thus tens of thousands of genes or RNAs can be simultaneously detected on a single array.
A use of hybridization reactions to identify the location of certain nucleic acids sequences were reported by several researchers. Gillespie discussed the idea of using support bound DNA to hybridize to complementary RNA1). Pardue and Gall2), and Jones and Robertson3) reported in situ hybridization, in which the positions of specific sequences can be located in the nucleus or chromosomes by hybridization reactions on cells fixed to microscopic slides. Later, the multicolor fluorescent labeling technique was developed for the analysis of multiple probes for in situ hybridation4, 5). In the late 1970s, Kafatos et al., introduced dot blots, a technique for analyzing multiple hybridization targets in parallel by applying them to a filter in a defined pattern6). In the dot blots, multiple targets are arrayed on the support and the probe, normally a single sequence, is labeled and applied under hybridization conditions to the membrane. Saiki et al. introduced a variant of dot blots, reverse dot blots, in which multiple probes are attached as an array to the membrane support and the target to be analyzed is labeled7).
The emergence of the Human Genome Project prompted various technological developments. One is the idea of using high-density oligonucleotide arrays for sequence analysis. Southern and Maskos reported in 1992 the synthesis of oligonucleotide arrays, which were a low-density prototype array8, 9). About one hundred different oligonucleotides up to 19 bases in length were synthesized in situ on a glass microscopic slide using parallel lines of silicon rubber tubing8). These oligonucleotide arrays were tested to examine the feasibility of using them for sequence analysis. Many of the basic ideas of DNA arrays such as arraying probes on solid supports and in situ preparation of probes, were already established by Maskos and Southern, and other researchers10), but how to prepare high-density microarrays was a completely different matter, requiring novel engineering intellect and knowledge, the photolithographic fabrication of the high-density oligonucleotide microarrays, and the robot spotting fabrication of DNA microarrays with pre-synthesized probes. |
|
|
