DNA Microarray Animation

A DNA microarray (also commonly known as gene or genome chip, DNA chip, or gene array) is a collection of microscopic DNA spots, commonly representing single genes, arrayed on a solid surface by covalent attachment to chemically suitable matrices. DNA arrays are different from other types of microarray only in that they either measure DNA or use DNA as part of its detection system. Qualitative or quantitative measurements with DNA microarrays utilize the selective nature of DNA-DNA or DNA-RNA hybridization under high-stringency conditions and fluorophore-based detection. DNA arrays are commonly used for expression profiling, i.e., monitoring expression levels of thousands of genes simultaneously, or for comparative genomic hybridization.
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Arrays of DNA can either be spatially arranged, as in the commonly known gene or genome chip, DNA chip, or gene array, or can be specific DNA sequences tagged or labelled such that they can be independently identified in solution. The traditional solid-phase array is a collection of microscopic DNA spots attached to a solid surface, such as glass, plastic or silicon chip. The affixed DNA segments are known as probes (although some sources will use different nomenclature such as reporters), thousands of which can be placed in known locations on a single DNA microarray. Microarray technology evolved from Southern blotting, whereby fragmented DNA is attached to a substrate and then probed with a known gene or fragment.





Applications of these arrays include:
  • mRNA or gene expression profiling - Monitoring expression levels for thousands of genes simultaneously is relevant to many areas of biology and medicine, such as studying treatments, disease, and developmental stages. For example, microarrays can be used to identify disease genes by comparing gene expression in diseased and normal cells .

  • comparative genomic hybridization (Array CGH) - Assessing large genomic rearrangements.

  • SNP detection arrays - Looking for Single nucleotide polymorphism in the genome of populations.
  • Chromatin immunoprecipitation (chIP) studies - Determining protein binding site occupancy throughout the genome, employing ChIP-on-chip technology.

Fabrication

Microarrays can be fabricated using a variety of technologies, including printing with fine-pointed pins onto glass slides, photolithography using pre-made masks, photolithography using dynamic micromirror devices, ink-jet printing, or electrochemistry on microelectrode arrays.


DNA microarrays can be used to detect RNAs that may or may not be translated into active proteins. Scientists refer to this kind of analysis as "expression analysis" or expression profiling. Since there can be tens of thousands of distinct probes on an array, each microarray experiment can accomplish the equivalent number of genetic tests in parallel. Arrays have therefore dramatically accelerated many types of investigations.


Spotted microarrays
In spotted microarrays (or two-channel or two-colour microarrays), the probes are oligonucleotides, cDNA or small fragments of PCR products that correspond to mRNAs and are spotted onto the microarray surface. This type of array is typically hybridized with cDNA from two samples to be compared (e.g. diseased tissue versus healthy tissue) that are labeled with two different fluorophores (e.g. Rhodamine (Cyanine 5, red) and Fluorescein (Cyanine 3, green)). The two samples are mixed and hybridized to a single microarray that is then scanned in a microarray scanner to visualize fluorescence of the two fluorophores. Relative intensities of each fluorophore are then used to identify up-regulated and down-regulated genes in ratio-based analysis. Absolute levels of gene expression cannot be determined in the two-colour array, but relative differences in expression among different spots (=genes) can be estimated with some oligonucleotide array.


Oligonucleotide microarrays

Two Affymetrix chips

In oligonucleotide microarrays (or single-channel microarrays), the probes are designed to match parts of the sequence of known or predicted mRNAs. T. These microarrays give estimations of the absolute value of gene expression and therefore the comparison of two conditions requires the use of two separate microarrays.

Oligonucleotide Arrays can be either produced by piezoelectric deposition with full length oligonucleotides or in-situ synthesis.

Long Oligonucleotide Arrays are composed of 60-mers, or 50-mers and are produced by ink-jet printing on a silica substrate. Short Oligonucleotide Arrays are composed of 25-mer or 30-mer and are produced by photolithographic synthesis on a silica substrate or piezoelectric deposition on an acrylamide matrix. More recently, Maskless Array Synthesis from NimbleGen Systems has combined flexibility with large numbers of probes. Arrays can contain up to 390,000 spots, from a custom array design. New array formats are being developed to study specific pathways or disease states for a systems biology approach.

Oligonucleotide microarrays often contain control probes designed to hybridize with RNA spike-ins. The degree of hybridization between the spike-ins and the control probes is used to normalize the hybridization measurements for the target probes.


Genotyping microarrays

DNA microarrays can also be used to read the sequence of a genome in particular positions.

SNP microarrays are a particular type of DNA microarrays that are used to identify genetic variation in individuals and across populations. Short oligonucleotide arrays can be used to identify the single nucleotide polymorphisms (SNPs) that are thought to be responsible for genetic variation and the source of susceptibility to genetically caused diseases. Generally termed genotyping applications, DNA microarrays may be used in this fashion for forensic applications, rapidly discovering or measuring genetic predisposition to disease, or identifying DNA-based drug candidates.

These SNP microarrays are also being used to profile somatic mutations in cancer, specifically loss of heterozygosity events and amplifications and deletions of regions of DNA. Amplifications and deletions can also be detected using comparative genomic hybridization, or aCGH, in conjunction with microarrays, but may be limited in detecting novel Copy Number Polymorphisms, or CNPs, by probe coverage.

Resequencing arrays have also been developed to sequence portions of the genome in individuals. These arrays may be used to evaluate germline mutations in individuals, or somatic mutations in cancers.

Genome tiling arrays include overlapping oligonucleotides designed to blanket an entire genomic region of interest. Many companies have successfully designed tiling arrays that cover whole human chromosomes.

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