Introduction Definition History Principle Types CDNA array Oligonucleotide array Microarray experiments Microarray fabrication Application Microarray database Conclusion Reference

1. By
KAUSHAL KUMAR SAHU
Assistant Professor (Ad Hoc)
Department of Biotechnology
Govt. Digvijay Autonomous P. G. College
Raj-Nandgaon ( C. G. )

2. SYNOPSIS
 Introduction
 Definition
 History
 Principle
 Types
 CDNA array
 Oligonucleotide array
 Microarray experiments
 Microarray fabrication
 Application
 Microarray database
 Conclusion
 Reference

3. Introduction
 It is understood that thousands of genes and their products (i.e., RNAs and proteins) in a given
living organism function in a complicated way that creates the mystery of life. However,
traditional methods in molecular biology generally work on a "one gene in one experiment"
basis, indicating that the throughput is very limited and the "global picture" of gene functions
is hard to obtain.
 In the past several years, a new technology, called microarray, has attracted tremendous
interests among scientists in biological research and other scientific fields. This technology
promises to monitor the whole genome on a single chip so that researchers can have a much
broader and better view of the interactions among thousands of genes simultaneously.
 The fundamental concept of microarray technology is to miniaturize traditional bio analytical
detection system so that hundreds or even thousands of biomolecules with unique identity can
be detected simultaneously in one single experiment by using a tiny amount of test sample.
 Microarray Technology is a powerful tool to monitor gene expression or gene expression
changes of hundreds or thousands of genes in a single experiment.

4. Definition
 An array is an orderly arrangement of samples. A DNA
microarray (also commonly known as DNA chip or biochip) is a
collection of microscopic DNA spots attached to a solid surface.
 monitoring expression levels for thousands of genes simultaneously.
Macroarrays contain sample spot sizes of about 300 microns or larger
and can be easily imaged by existing gel and blot scanners.
The sample spot sizes in microarray are typically less than 200
microns in diameter and these arrays usually contain thousands of
spots.

5. History
 Microarray technique was first discovered by Mark
Schena et al in 1995.
 The first complete eukaryotic genome (Saccharomyces
cerevisiae) on microarray was published in 1997.
 Microarray technology evolved from Southern blotting,
where fragmented DNA is attached to a substrate and then
probed with a known gene or fragment.

6. Principle
 The underlying principle of microarray
technology is base-pairing or
hybridization (A-T and G-C) the
ability of DNA to bind to itself and to
RNA.
Term definitions
 TARGET: cDNA or oligonucleotide or
single-stranded DNA with a defined
identity attached to the array surface
 PROBE: the labeled DNA or RNA
within the hybridization mix (which
can hybridize to the complementary
probe strand)

7. Types of microarray
1.cDNA arrays
 A DNA molecule derived from mRNA with the help of enzyme reverse
transcriptase is termed as cDNA or complementary DNA. The cDNA fragments
(500-2500 bp) are spotted on a suitable substrate and hybridized with labeled targets
for gene expression studies. The probe sequences have been taken straight from
sequence database (UniGene, GeneBank etc.)
2.Oligonucleotide arrays
 Contrary to cDNA microarrays, the oligonucleotide microarrays are made by
chemical synthesis using computer algorithms, each oligonucleotide (15-70
nucleotides long) represents a part of known gene.
Two Main Types of Microarrays
cDNA arrays: spotted onto surface
Oligonucleotide arrays: created on surface

8. Microarray experiments
 A typical microarray experiment
consists of
 extracting RNA from the cells or tissue
being examined,
 converting the RNA to cDNA, by RT
enzyme and amplify by PCR,
 labelling the cDNA with fluorescent
dyes and
 allowing the labelled cDNA to
hybridize with the material (cDNA or
oligonucleotide) on the microarray slide.

9. cDNA Arrays -experiments

10. Oligonucleotide versus cDNA Arrays-
 cDNA arrays can be used
for 2 samples analysis $
oligonucleotied arrays are
used only for 1 sample
analysis.
 cDNA arrays does not
require probe designing.
 cDNA arrays provides
higher specificity due to
longer sequences of
targets.
 cDNA array more useful
on a global level

11. Microarray Fabrication
Two different approaches are used for microarray production
 Presynthesized probes with subsequent deposition on chip, and
 In situ (on chip) synthesis of probes.
The presynthesized probes, i.e., whole
genes, small sequences or products of PCR
are prepared independently without any
involvement of chip.
 Then The robot automatically collects
samples (probes) from 96 or 384 well
microtiter plates into its thin pins or pens
and carries them to the chip surface for
deposition.
 After deposition of samples, the chip is air
dried and exposed to ultraviolet radiation so
that samples are immobilized due to
covalent bonding between sample
molecules and chemical compound
(polylysine, amino silanes or amino reactive
silanes) coated onto chips.
 Finally, the deposited samples are
rendered single stranded by heat or alkali
treatment.
Presynthesized probes with subsequent deposition on chip
Fig:ROBOT SPOTTER

12. In situ (on chip) synthesis of probes
The advantage of using impermeable surface like glass
allows in situ synthesis of pobes using combinatorial
strategies for the fabrication of microarray in a few
coupling steps. The yields of in situ nucleic acid
synthesis are consistent and high.
 Some methods for direct synthesis of oligonucleotide
on defined areas on a chip are:
 Ink-jet printing
 Photolithography
 Electrochemical synthesis

13. InkJet (HP/Canon) technology

14. Febit/NimbleGen photolitography

15. Application of Microarray:-
1. Gene discovery: DNA Microarray technology helps in the
identification of new genes, know about their functioning and
expression levels under different conditions.
2. Disease diagnosis: DNA Microarray technology helps researchers
learn more about different diseases such as heart diseases, mental
illness, infectious disease and especially the study of cancer.
3. Drug discovery: 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.

16. Application of Microarray:-
4. Toxicological research: 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.
5. Gene expression profiling: In an mRNA or gene expression profiling
experiment the expression levels of thousands of genes are simultaneously
monitored to study the effects of certain treatments, diseases, and
developmental stages on gene expression.
6. Comparative genomic hybridization: Assessing genome content in
different cells or closely related organisms.
7. SNPs analysis: can be easily detected by providing highly stringent
hybridization conditions during microarray experiment. Under these
condition a single base mismatch leads to unhydridized target-prode.

17. Microarray Databases
 The term microarray database is usually used to
describe a repository containing microarray gene
expression data. The key features of a microarray
database are to store the measurement data, manage a
searchable index, and make the data available to other
applications for analysis and interpretationThere are
several databases available online which serve as
repositories of microarray

18. GEO at the NCBI
 The gene expression
omnibus (GEO) is a
leading public repository
 that stores and distributes
high throughput gene
expression data generated
by scientific community
from a wide range of
biological issues.
 Data can be searched
either from the gene or
experiment perspectives.
 In addition, several user
friendly tools are
available for the analysis
of microarray data.

19. Array express
 Array express is a
public resource for
microarray data,
which stores well-
annoted raw an
normalized data.
 These data are
stored in the data
warehouse and can
be queried on gene,
sample, and
experiment
attributes.
 Users are allowed
to browse and query
on experiment
properties, array
platform, species,
authors, etc.

20. Conclusion
 Microarray is a revolutionary technology and microarray
experiments produce considerably more data than other
techniques. Integrating gene expression data with other
biomedical resources will provide new mechanistic or
biological hypotheses. However, innovative statistical
techniques and computing software are essential for the
successful analysis of microarray data.