DNA microarray

DNA microarray is like a microscope slide made of glass, silicon

or nylon. It is also known as DNA chip or biochip. Each spot has
picomoles of a specific DNA sequence, known as probes (or
reporters or oligos). These are short sections of a gene or other
DNA that hybridize a cDNA or cRNA.

The “probe-target” hybridization is detected and quantified

using fluorescence to determine relative abundance of a nucleic
acid sequence in the target. The total strength of the signal,
from a spot (feature), depends on the amount of target sample
binding to the probes present on that spot.

The DNA that is to be tested is prepared from the mRNA by the

action of reverse transcriptase as cDNA. [ against the normal
gene sequence as control]. The cDNAs are linked to a
fluorescent dye. [ test sample sequence with red and normal
gene sequence as control with green]. The DNA probes of the
microarray are exposed to these cDNAs. Any complementary
sequences present on the cDNAs will bind to the fixed probes
After this. the microarray is rinsed free of any cDNA that has not
hybridised. Then it is exposed to laser light that causes fluorescence where hybridization occurred.
The brighter the light emitted, the higher the level of hybridization and in turn higher the expression
of that gene in the cell.

A full human genome microarray would have as many as 30,000 spots. Others are made as clusters
of genes that relate to particular conditions, such as cancers.

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.

For example, microarray-based gene expression profiling can be used to identify genes whose
expression is changed in response to pathogens or other organisms by comparing gene expression in
infected to that in uninfected cells or tissues

DNA Probe :
DNA Probe is a single stranded, 10 to 10,000bp long DNA complementary to the specific base
sequence of a DNA fragment of interest. DNA probe comprises all or most of the bases of a target
gene. It is synthesised chemically.
DNA Microarray Procedure

To compare Gene Expressions of two samples [test and normal control]

1. Obtain mRNAs from test and normal control organisms [disease affected cell and a healthy
cell].
2. Reverse transcription of mRNAs to produce cDNAs for both [as ssDNA strand]

3. Add different fluorescent labels to cDNAs of test [red] and normal control [green]
4. Mix the fluorescent labeled cDNAs of test [red] and normal control [green]

5. Apply mixed cDNAs on the Microarray that contains ssDNA probes of various genes of the
species being tested.

6. cDNA hybridises with ssDNA probes on microarray

7. After washing, scan the Microarray using laser scanning to record fluorescence pattern
8. Fluorescence shows the expressed genes. [ Presence of mRNA, in this case the cDNA,
indicates expression of gene]. And intensity of fluorescence shows the level of gene
expression
9. Compare the fluorescence, in turn the Gene Expression, between test and normal control