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INTRODUTION
A DNA microarray is a multiplex technology used in molecular biology and in medicine.
Microarray technology evolved from southern blotting where fragmented DNA is attached
to a subtrateand then probed with a known gene or fragment .
The basic principle underlying microarray technology is that complementary nucleic acids
will hybridize. Hybridization provides exquisite selectivity of complementary stranded nucleic
acids, with high sensitivity and specificity. In the traditional techniques, in which radioactive
labeling materials are usually used, the simultaneous hybridization of test and reference samples
is impossible
DNA microarray technology has empowered the scientific community to understanding the
fundamental aspect underlying the growth and development of life as well as to explore the
genetic cause of anomalities occurring in the functioning of the human body.
A typical microarray experiment involves the hybridization of an mRNA molecule to the
DNA template from which it is originated.
The advantages of the cDNA microarray compared with the oligonucleotide array have been
thought to include less susceptibility and higher specificity due to the longer sequences of the
targets .However, cDNA may contain repetitive sequences that are often observed in various
genes, or similar sequences that are found in family member genes. These non-specific sequences
may affect the sensitivity of the cDNA microarray.
Types of microarray
Depending upon the kind of immobilized sample used construct arrays and the information
fetched, the Microarray experiments can be categorized in three ways:
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1. Microarray expression analysis
2. Microarray for mutation analysis
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3. Comparative Genomic Hybridization
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Principal of micro array technology;
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The basic principle underlying micro array technology is that complementary nucleic acids will
hybridize. Hybridization provides exquisite selectivity of complementary stranded nucleic acids,
with high sensitivity and specificity. In the traditional techniques, in which radioactive labeling
materials are usually used, the simultaneous hybridization of test and reference samples is
impossible
In micro array-based technologies, the solid surface, such as a glass slide, contains hundreds
to thousands of immobilized DNA (targets) spots which can be simultaneously hybridized with
two samples (probes) labeled with different fluorescent
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Applications include:
Technology or
Synopsis
Application
used to identify genes whose expression is changed in response
Gene expression
to pathogens or other organisms by comparing gene expression
profiling
in infected to that in uninfected cells or tissues.
DNA sequences bound to a particular protein can be isolated
Chromatin
by immunoprecipitating that protein these fragments can be
immunoprecipitation on
then hybridized to a microarray allowing the determination of
Chip
protein binding site occupancy throughout the genome.
microarrays make use of SNP detection, including Genotyping,
forensic analysis, measuring predisposition to disease,
SNP detection identifying drug-candidates, evaluating germline mutations in
individuals or somatic mutations in cancers, assessing loss of
heterozygosity, or genetic linkage analysis.
Gene expression:
The sequence of a gene does not give information about its activity. A gene must be
transcribed into mRNA to be expressed (or turned ON). The level of expression (or activity) of a
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gene can therefore be studied by measuring how much corresponding mRNA is made
(transcribed) from a particular gene.
The sequence of a gene is complementary to the sequence of its mRNA. Therefore the mRNA
can base pair (or hybridize) to the DNA strand it was copied from (template strand). Scientists
study gene expression by measuring the amount of mRNA for different genes.
ALL of our cells contain the exact same DNA sequence. All cells arose from a single cell (the
egg). However different type of cells in the body expresses different sets of genes. For example,
a liver cell expresses some of the same genes expressed in lung cells or skin cells but also
expresses genes that no other cell type expresses (liver-specific genes). As a result liver cells
look different from other cells in the body and do things that no other cell can do.
Microarrays are simply a method for visualizing which genes are likely to be used in a
particular tissue at a particular time under a particular set of conditions. The output of a
microarray experiment is called a “gene expression profile.”
Flow Diagram of Gene Expression Profiling
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Drug Target:
DNA microarray which allows the monitoring of gene expression level of thousands of genes
simultaneously played a significant role in drug discovery.
Animals may not be fully predictive of the response in human due to species variation. The
pre-clinical studies in animals do not always detect drugs that prove to be toxic in human. Ex.
tacrine, used to treat patients with Alzheimer’s disease include hepatotoxicity by the elevation of
liver transminase in human, but there was no evidence of liver toxicity in mice rats or dogs
during pre-clinical development. Drug-induced hepatotoxicity and liver damage cause significant
morbidity and mortality and represent a major challenge in the area of drug development and
design.
Microarray databases are built on the expression data from well defined marketed
pharmaceuticals, classical chemical toxicants and few proprietary drugs. These compounds have
well defined pharmacological action and some are rat and human specific, which result in data
bases that are useful for predicting toxicity during development of toxicity.
Microarray can be used to screen for changes in gene expression following exposure of
tumour cells to drugs either in culture or in patients following treatment. Studies on this type
may provide information on the precise mechanism of action of the drug or could lead to the
identification of early markers of drug response. For predicting the longer –term clinical
response to drug treatment, microarry could also yield information about possible side effects,
and identify markers that, in clinical context, could be used to predicts possible adverse effect
events.
Application of Microaray:
Use of Microarrays in Tissue Engineering, Phenotype Analysis, and Monitoring
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Microarray Application to Oral Biology and Medicine
Gene discovery
Drug discovery and development
Toxicological research
Tumour classification
Tumour growgh and progression
Disease diagnosis
Detection, identification, and characterization of food-borne pathogens
DNA microarrays are being analyzed to understand the innate immune system, including
the cellular response to various bacterial endotoxins
DNA microarrays are now being used to identify other genes that may be regulated in
response to endotoxins, such as LPS, with the ultimate hope of providing new drug
targets for sepsis With the use of human umbilical vein endothelial cells as a model,
investigators found various genes that appear to be regulated in response to LPS. Many
of the genes identified confirm already well-established LPS-regulated genes, whereas a
few may represent new gene targets
Microarray technology has also been used to monitor changes in metabolic processes
including aging
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Challenges:
The limited availability or complete absence of in vivo human data adds additional
challenges to the analysis of microarrays using comparative genomics. In such an instance, the
effects observed using in vivo animal models and in vitro human data need to be projected to the
effects in humans. Hence, a challenge is to extrapolate from these limited studies whole-body
human effects while accounting for potentially not well-known or wellunderstood nonlinearities.
Lack of Appropriate Arrays
Lack of Comparable Studies
Applications will encompass defining gene function;
Inferring functional networks and pathways;
Understanding how variation is distributed among individuals, populations, and species;
Developing clinical protocols relating to cancer prognosis and detection of toxin
exposure
Conclusions:
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The power of microarray technology is that it allows global analysis of patterns of
simultaneous gene expression.
Microarry expression patterns could be integrated on the one hand with clinical data to
identify new markers to predict biological behavior of tumors and on the other hand with
database of drug sensitivity to unravel the molecular basis of drug action.
REFERENCES
Abiko Y, Hiratsuka K, Ohta M, Sasahara S, Tsujimoto A, Ohashi N (2000). Gene-
microarray analysis of gene expression in gingival epithelial cells by LPS
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Adams JC, Watt FM (1993). Regulation of development and differentiation by the
extracellular matrix. Aharoni A, Keizer LCP, Bouwmeester HJ, Sun Z, Huerta MA,
Verhoeven HA, et al. (2000). Identification of the SAAT gene involved in strawberry
flavor biogenesis by use of DNA microarrays.
Alizadeh AA, Eisen MB, Davis ER, Ma C, Lossos IS, Rosenwald A, et al. (2000).
Distinct types of diffuse large B-cell lymphoma identified by gene expression profiling.
Ashkenas J, Muschler J, Bissell MJ (1996). The extracellular matrix in epithelial biology:
shared molecules and common themes in distant phyla.
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