2. DNA MICRO ARRAY
A DNA microarray (also commonly known as DNA chip or biochip) is a
collection of microscopic DNA spots attached to a solid surface. Scientists
use DNA microarrays to measure the expression levels of large numbers
of genes simultaneously or to genotype multiple regions of a genome.
3. Introduction.
The large scale genome sequencing effort and the ability to immobilize
thousands of DNA fragments on coated glass slide or membrane, have led
to the development of micro array technology
A microarray is a pattern of ssDNA probes which are immobilized on a
surface called a chip or a slide.
Microarrays use hybridization to detect a specific DNA or RNA in a sample.
It uses million different probes fixed on a solid surface.
4. PRINCIPLE
The core principle behind
microarrays is hybridization
between two DNA strands.
Fluorescent labeled target
sequence that binds to a probe
sequence generated signal that
depends on the strength of the
hybridization determined by the
number of paired bases
5. PRINCIPLE
The principle is based on the fact that complimentary sequence of DNA
can be used to hybridize, immobilized DNA molecules.
There are five major steps in performing a microarray.
1. Sample preparation and labelling
2. Hybridization
3. Washing
4. Image acquisition
5. Data analysis
6. The Workflow of Microarray
Array
Hybridized Array
Hybridization
Scanning
Plate
Array Fabrication
Plate Preparation
RNA extraction
Labeled cDNA
cDNA synthesis
and labeled
sample
7. Sample preparation and
labeling
• Isolate a total RNA containing mRNA, that ideally represents
a quantitative copyof genes expressed at the time of sample
collection.
• Preparation of cDNA from mRNA using a reverse
transcriptase enzyme.
• Short primer is required to initiate cDNA synthesis
• Each cDNA is labelled with fluorescent dyes for e.g.. Cyanine
dyes(i.e Cy3 & Cy5)
8. Array in hybridization
Here the labelled cDNA
sample are mixed together.
Purification
After purification, the mixed
lablled cDNA sample is
completely hybridized
against denatured PCR
product or cDNA molecules
spotted on a glass slide.
9. Image acquisition and data
analysis
• Slide is dried and scanned to
determine how much
labelled cDNA (probe) is
bound to each target spot.
• Hybridize target produces
emissions
• Microarrays software often
uses green, red and yellow
spots on the microarray to
represents hybridization.
10. Applications :
1. Microarray as a gene expression profiling tool
2. Microarray as a comparative genomics tool
3. Disease diagnosis
4. Drug discovery
5. Toxicological research
11. MICROARRAY AS A GENE
EXPRESSION PROFILING TOOL
1.The principle aim of using microarray technology as a gene
expression profiling tool is to answer some of the
fundamental questions in biology, such as “When”, “Where”
and “What” magnitude genes of interest are expressed.
2. Microarray analysis measures changes in the multigene
patterns of expression to better understand about
mechanisms and broader bioactivity functions of gene.
12. Microarray as a comparative genomics
tool
1. Microarray technology have widespread use in comparative gene
mutation analysis to analyze genomic alterations such as sequence and
single nucleotide polymorphisms.
In microbiology microarray, gene mutation analysis is directed to
characterization of genetic differences among microbial isolates,
particularly closely related species.
13. Disease diagnosis
Different types of cancers have been classified on the basis of the
organs in which the tumors develop.
Now,with the evolution of microarray technology, it will be possible for
the researches to further classify the types of cancer on the basis of
patterns of gene activity in tumor cells.
14. TOXICOLOGICAL RESEARCH
1.Microarray technology provides a platform for the research of the impact of
toxins on the cells and their passing onto the progeny.
2. Toxicogenomics establishes correlation between responses to toxicants and
the changes in the genetic profiles of the cells exposed to such toxicants.
3. The microarray permits researchers to examine thousand of different genes
in the same experiments and thus to obtain a good understanding of the
relative levels of the expressions between different genes in an organism.
