A DNA microarray contains thousands of single-stranded DNA probes arranged in a grid on a glass or silicon chip. The DNA of interest is fluorescently labeled and can hybridize to its complementary probe on the microarray.

This technique allows the simultaneous measurement of the relative concentrations of specific cDNA sequences.

Commonly, DNA microarrays are used to quantify gene expression by measuring the levels of cDNA corresponding to particular mRNA transcripts.

Other uses include detecting the binding of transcription factors on chromatin or identifying single-nucleotide variations in genomic DNA.

Microarray probes for measuring mRNA are single stranded DNA segments corresponding to each gene’s coding region to be analyzed. The probe nucleotides can either be copied from genomic DNA using PCR, or synthesized directly onto the chip’s surface.

These probes are covalently attached to a chip forming an array of spots, where each site represents a single target. Therefore, one DNA microchip can analyze tens of thousands of genes.

For a genome-wide analysis, the total mRNA content is isolated from the cells of an organism and converted to fluorescently labeled cDNAs. When this sample is exposed to a DNA microarray chip, the probes pair with their complementary cDNAs.

The fluorescent signal at each spot is directly proportional to the amount of cDNA bound, and can be analyzed by the appropriate software to quantify gene expression.

Although this analysis does not yield the absolute concentrations of the mRNAs, it can be used to compare their relative concentrations— hence gene expression levels, in two samples under different conditions.

A reference sample could be labeled with a green fluorophore, while a test sample would be labeled red.

If the test sample has a lower expression than the reference sample, then the spot appears green, while a higher expression is indicated by a red spot. Yellow indicates equal expression levels in both samples.

Microarrays are high-throughput and relatively inexpensive assays that can be automated to analyze large quantities of data at a time. They are used in genome-wide studies to compare gene or protein expression under two varied conditions, such as healthy and diseased states. Microarrays consist of glass or silica slides on which probe molecules are covalently attached through surface functionalization. Most commonly, the slides are prepared through the chemisorption of silanes to silica surfaces.

While DNA microarrays are used to monitor gene expression, protein microarrays detect biologically relevant interactions of a fluorescently labeled protein with other proteins, DNA, or small molecules.

DNA microarray slides are often called gene chips or DNA chips. DNA chips are widely used to profile thousands of genes parallelly. Other than gene expression, they are also used in studying microRNA expression, DNA copy numbers, and single-nucleotide polymorphism detection. Commercially, these chips are used in biotechnology, forensics, toxicology, drug discovery, and targeted therapy for infectious and genetic diseases.

The chemically stable nature of DNA probes makes DNA chips the most popular of the microarray technologies. The covalently bound single-stranded DNA probe retains its native interactions despite harsh chemicals and temperatures that are used during probe loading and chip preparation.

The most obvious shortcoming of DNA microarrays is the prerequisite of a known target sequence. Because DNA microarray probes are designed to complement the target cDNA sequence, the sequence of the mRNA from which the cDNA is copied must be known. This makes the technology unusable while looking for novel mRNA sequences. For such purposes, RNASeq is the preferred albeit expensive option.

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