JAIPUR NATIONAL UNIVERSITY

PRESENTED BY: RICHA KUMARI

BRANCH : B.TECH BIOTECH 6TH SEM
1. INTRODUCTION
2. HISTORY
3. PRINCIPLE
4. DNA MICROARRAY TECHNOLOGY
5. PRINCIPLES OF DNA MICROARRAY TECHNOLOGY
6. TYPES OF DNA MICROARRAY
GLASS cDNA MICROARRAYS
IN SITU OLIGONUCLEOTIDE ARRAY FORMAT
7. APPLICATIONS OF MICROARRAY TECHNOLOGY
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
microarray 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.
DNA microarray uses a million different probes, fixed on
a solid surface.
 An array is an orderly
arrangement of samples where
matching of known and
unknown DNA samples is done
based on base pairing rules.

An array experiment makes use

of common assay systems such
as microplates or standard
blotting membranes.

Fig-01 Robotic arm with

spotting slides
 Microarray technology evolved from Southern
blotting.
The concept of microarrays was first proposed in the
late 1980s by Augenlicht and his colleagues.
They spotted cDNA sequences on nitrocellulose
membrane and used radioactive labeling to analyze
differences in gene expression patterns among
different types of colon tumors in various stages of
malignancy.
 The core principle behind
microarrays is hybridization
between two DNA strands.

Fluorescent labeled target

sequences that bind to a probe
sequence generate a signal that
depends on the strength of the
hybridization determined by
the number of paired bases.

Fig-02 Array hybridization

 DNA microarray technology may be defined as a
high-throughput and versatile technology used for
parallel gene expression analysis for thousands of
genes of known and unknown functions.
Used for detection of polymorphisms and mutations
in genomic DNA
A DNA microarray is a collection of microscopic
DNA spots on solid surface. Each spot contains
picomoles of a specific DNA sequence, known as
probes or reporters.
 Each identified sequenced gene on the glass, silicon
chips or nylon membrane corresponds to a fragment
of genomic DNA, cDNAs, PCR products or
chemically synthesized oligonucleotides of up to
70mers and represents a single gene.

Probe-target hybridization is usually detected and

quantified by detection of fluorophore, silver, or
chemiluminescence labeled targets to determine
relative abundance of nucleic acid sequences in the
target.
 The principle of DNA microarray technology is based
on the fact that complementary sequences of DNA
can be used to hybridize , immobilized DNA
molecules.
There are four major steps in performing a typical
microarray experiment.
Sample preparation Image acquisition
and Hybridization Washing and
labeling Data analysis
 Isolate a total RNA containing
mRNA that ideally represents a
quantitative copy of 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 (Sample and Control)
is labelled with fluorescent
cyanine dyes (i.e. Cy3 and Cy5). Fig-03 Sample labeling
 Here, the labelled cDNA
(Sample and Control) are
mixed together.
Purification
After purification, the
mixed labelled cDNA is
competitively hybridized
against denatured PCR
product or cDNA molecules
spotted on a glass slide.

Fig-04 Array Hybridisation

 Slide is dried and scanned to
determine how much labelled
cDNA (probe) is bound to
each target spot.
Hybridized target produces
emissions.
Microarray software often uses
green spots on the microarray
to represent upregulated genes.
Red to represent those genes
that are downregulated and
yellow to present in equal
abundance
Fig-05 Gene chip showing different
type of color spots
1) Glass cDNA microarrays which involves the micro
spotting of pre-fabricated cDNA fragments on a glass
slide.

2) High-density oligonucleotide microarrays often

referred to as a "chip" which involves in
situ oligonucleotide synthesis.
 Glass cDNA microarrays
was the first type of DNA
microarray technology
developed.
It was pioneered by Patrick
Brown and his colleagues at
Stanford University.
Produced by using a robotic
device which deposits
(spots) a nanoliter of DNA
onto a coated microscopic
glass slide (50-150 m in
diameter) .
Fig-06 Contact printer with
robotic pins
Selection of the material to
spot onto the microscope glass
surface.

Preparation and purification of

DNA sequences representing
the gene of interest.

Spotting DNA solution onto

chemically modified glass
slides via a contact printing or
inkjet printing.
FIG-07 Spotting of
slides
 Advantages of Glass cDNA microarrays include their
relative affordability with a lower cost.
Its accessibility requiring no specific equipment for use
such that hybridisation does not need specialised
equipment.
Data capture can be carried out using equipment that is
very often already available in the laboratory.
 Glass cDNA microarray have a few disadvantages such
as intensive labour requirement for synthesizing,
purifying, and storing DNA solutions before microarray
fabrication.
They may hybridise to spots designed to detect
transcript from a different gene.
MICROARRA MICROARRA
Y AS A GENE Y AS A DISEASE DRUG TOXICOLOGIC
EXPRESSION COMPARATIV
DIAGNOSIS DISCOVERY AL RESEARCH
PROFILING E GENOMICS
TOOL TOOL
 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 to what
magnitude genes of interest are expressed.

Microarray analysis measure changes in the multigene patterns

of expression to better understand about regulatory
mechanisms and broader bioactivity functions of genes.
 Microarray technology have widespread use in
comparative gene mutation analysis to analyse
genomic alterations such as sequence and single
nucleotide polymorphisms.

In microbiology microarray gene mutation analysis is

directed to characterisation of genetic differences
among microbial isolates, particularly closely related
species.
 Different types of cancer 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 researchers to further classify
the types of cancer on the basis of the patterns of gene
activity in the tumor cells.
 Microarray technology has extensive application in
Pharmacogenomics.

Comparative analysis of the genes from a diseased and

a normal cell will help the identification of the
biochemical constitution of the proteins synthesized by
the diseased genes.
 Microarray technology provides a robust platform for the
research of the impact of toxins on the cells and their passing
on to the progeny.
Toxicogenomics establishes correlation between responses to
toxicants and the changes in the genetic profiles of the cells
exposed to such toxicants.
The microarray permits researchers to examine thousands of
different genes in the same experiment and thus to obtain a
good understanding of the relative levels of expression
between different genes in an organism.
 Microarray is a recently developed functional genomics
technology that has powerful applications in a wide
array of biological medical sciences, agriculture,
biotechnology and environmental studies. Since many
universities research institutions and industries have
established microarray based core facilities and
services, microarrays have become a readily accessible,
widely used technology for investigating biological
systems.
 https://www.ncbi.nlm.nih.gov/pmc/articles/P
MC4011503/
http://learn.genetics.utah.edu/content/labs/
microarray/
http://www.premierbiosoft.com/tech_notes/
microarray.html
THANK YOU