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Current Knowledge on Microarray Technology - an Overview Naidu & Suneetha Tropical Journal of Pharmaceutical Research February 2012; 11 (1): 153-164 © Pharmacotherapy Group, Faculty of Pharmacy, University of Benin, Benin City, 300001 Nigeria. All rights reserved . Available online at http://www.tjpr.org http://dx.doi.org/10.4314/tjpr.v11i1.20 Review Article Current Knowledge on Microarray Technology - An Overview * Chitrala Kumaraswamy Naidu and Yeguvapalli Suneetha Department of Zoology, Sri Venkateswara University, Tirupati -517502, India. Abstract The completion of whole genome sequencing projects has led to a rapid increase in the availability of genetic information. In the field of transcriptomics, the emergence of microarray-based technologies and the design of DNA biochips allow high-throughput studies of RNA expression in cell and tissue at a given moment. It has emerged as one of the most important technology in the field of molecular biology and transcriptomics. Arrays of oligonucleotide or DNA sequences are being used for genome-wide genotyping and expression profiling, and several potential clinical applications have begun to emerge as our understanding of these techniques and the data they generate improves. From its emergence to date, several database, software and technology updates have been developed in the field of microarray technology. This paper reviews basics and updates of each microarray technology and serves to introduce newly compiled resources that will provide specialist information in this area. Keywords: Microarray, Databases, cDNA array, Oligonucleotide array Received: 19 April 2011 Revised accepted: 19 December 2011 *Corresponding author: Email: [email protected]; Tel: +91 9985563644 Trop J Pharm Res, February2012;11 (1): 153 Naidu & Suneetha INTRODUCTION or radioactive tag and a brighter signal is detected when more copies bind [4]. In The completion of whole genome sequencing general, all microarray assays contain five projects has led to a rapid increase in the discrete experimental steps – biological availability of genetic information. In the field query, sample preparation, biochemical of transcriptomics, the emergence of reaction, detection, data visualization and microarray-based technologies and the modeling [5]. According to Yuk Fai leung et al design of DNA biochips allow high-throughput [6] , a typical microarray experiment involves studies of RNA expression in cell and tissue sample extraction, fluorescent labeling, co- at a given moment [1]. Today, microarray hybridization, scanning and finally statistical technology is one of the popular tools in analysis. Microarray technologies can be molecular biology with its main advantage broadly categorized into DNA microarrays being that, unlike other traditional methods, it and protein microarrays. is not limited to investigating ‘one gene at a time’ [2]. DNA microarrays In this review, we explain some of the DNA microarrays provide a simple and fundamentals and recent updates on each natural vehicle for exploring the genome in a microarray technology step-by-step. As it will way that is both systematic and be overwhelming to discuss all the updates of comprehensive. The power and universality microarrays since its emergence, we have of DNA microarrays as experimental tools focused on the updates of the past few years. derives from the exquisite specificity and We intend also to expound on current affinity of complementary base-pairing [7]. knowledge of recent databases, data analysis DNA microarrays are of oligonucleotide software and some of the companies in the arrays and a variety of cDNA arrays. field of microarrays. We hope that an understanding of some of the fundamentals, Oligonucleotide arrays recent updates and commercial technologies in microarrays will afford the beginner useful Oligonucleotide arrays or DNA chips are information in this area. miniature parallel analytical devices containing libraries of oligonucleotides MICROARRAY TECHNOLOGY robotically spotted (printed) or synthesized in situ on solid supports (glass, coated glass, Microarray is a technology which allows silicon or plastic) in a such way that the quantitative, simultaneous monitoring and identity of each oligonucleotide is defined by expression of thousands of genes [3]. The its location [8]. Oligonucleotide arrays contain basic principle behind microarray is the base short fragments of DNA (25 base pairs). One complementarity, i.e., the base pair ‘A’ is of the commercially available oligonucleotide complementary to ‘T’ and ‘C’ is microarrays are the Gene Chips developed complementary to ‘G’. In a microarray, many by firms such as Affymetrix. Ten to hundred thousands of spots are placed on a thousands of different oligonucleotide probes rectangular grid with each spot containing a are synthesized on each array. Traditionally, large number of pieces of DNA from a Affymetrix GeneChip Arrays are particular gene. When the sample of interest manufactured as a single array caged in a contains many copies of mRNA, many sealed cartridge with glass as a substrate. bindings will occur, indicating that the gene from the transcribed mRNA is highly The oligonucleotide technology pioneered by expressed. The quantity of hybridization can Affymetrix GeneChips differs from cDNA be determined because each copy of mRNA microarray in two important respects [9]. is labeled in the experiment with a fluorescent First, the probes are a set of 20-25 short Trop J Pharm Res, February2012;11 (1): 154 Naidu & Suneetha oligonucleotides that are specific for each onto a solid support (glass slide or gene or exon, along with the related set with membrane). Some of the basic principles single base mismatches incorporated at the behind preparation of cDNA arrays includes: middle position of each oligonucleotide. i) selection of the targets to be printed on the These are synthesized in situ on each silicon array directly from databases such as chip using genome sequence information to GenBank, dbESt, and UniGene or randomly guide photolithographic deposition. Second, from any library of interest; ii) arraying the the arrays are hybridized to a single selected cDNA targets onto the known biotinylated amplified RNA sample, and the location of coated glass microscope slide intensity measure for each gene is computed using a computer-controlled high speed by an algorithm that messages the difference robot; iii) fluorescently labeling the total RNA between the match and mismatch from both test and reference samples using measurements, and averages over each dyes with a single round of reverse oligonucleotide. Key difference between gene transcription; iv) pooling the florescent target chips and a cDNA microarray is the way for hybridization under stringent conditions; v) genes are represented on the arrays. In in measuring the laser excited incorporated situ photolithographic synthesis method used targets using a scanning confocal laser by Affymetrix gene chip probe arrays, the microscope; and v) finally, analyzing the quality of chips produced depends critically images from scanner by importing into a on the efficiency of photo-deprotection [10]. software in which they are pseudo-colored and merged. Micro spotting, piezoelectric There are many other advantages to printing and photolithography (an ‘ in situ ’ oligonucleotide-based microarrays. The fabrication technique developed by quality and reproducibility of printed Affymetrix) are some of the techniques used oligonucleotide arrays (“print format”) are for arraying cDNA. Invitrogen, Genome superior [11]. Although there is a large initial systems, Biodiscovery, Xeno, Affymetrix, capital outlay to purchase large sets of Silicon genetics, Genetix, etc., are some of oligonucleotides, the printed arrays are the companies which produce commercial inexpensive on a unit basis. Furthermore, arrayers. Open GMS 418 Array Scanner, long oligonucleotide probes can be GeneChip® Scanner 3000 System synthesized directly on an array surface (Affymetrix), DNA Microarray Scanner (photo printed arrays). In either case, whether (Agilent), Typhoon Variable Mode Imager printed or photo printed, the composition of (GE Healthcare), InnoScan 700( INNOPSIS), the array can be absolutely specified and Axon GenePix4000ALMicroarray scanner hence is completely reproducible by others. A (Molecular Devices), and PowerScanner limitation of oligonucleotide arrays is the (Tecan) are some of the commercial requirement of relatively large amounts of scanning devices which detect different levels biological starting materials for gene of fluorescence between the spots on the expression analysis [12]. GE Healthcare, microarray. Apart from applications of cDNA Ocimum Biosolutions, and Agilent are some array in comparative genome analysis and of the other companies which provide functional genome analysis, recent commercially available designs of applications include gene expression profiling Oligonucleotide microarrays. of drug-resistant small cell lung cancer cells [13], characterization of osmotic stress- cDNA arrays induced genes in poplar cells [14] and veterinary diagnostics. cDNA arrays contain long fragments of DNA (from 100 to thousands of base pairs). cDNA arrays are created by robotically spotting Protein microarray individual samples of purified cDNA clones Trop J Pharm Res, February2012;11 (1): 155 Naidu & Suneetha Though DNA microarray is very useful, it has used ones. In antibody microarrays, bait several limitations. The expression levels of molecules are numerous individual antibodies many genes are subject
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