MICROARRAY

Microarray gene expression

To understand microarray gene expression, we will consider the example of an apple tree throughout our discussion.

                                               An apple tree

The example of an apple tree

The apple tree genome consists of thousands of genes. In any one tissue only some of the genes are expressed. Each cell within the leaf contains entire apple genome. The same is true for apple flower and fruit. But the set of genes that are expressed in the leaf may or may not be the same as those expressed in the flower and the fruit. Or it may also be that although same genes are being expressed, the level of expression varies.

Healthy apple vrs infected apple

We may also compare the expression of genes in different samples of the same tissue, for example the gene expression pattern in a healthy apple and an infected apple.

Genomics and Microarray

In genomics the goal is to identify genes expressed in each tissue. Microarray is a tool that allows study of gene expression in many tissues simultaneously

Microarray

Microarrays are prepared in a glass slide. SS DNA fragments from each gene in the genome are spotted in glass slide. DNA spots are arranged in specific order. RNA is extracted from a particular tissue. Complementary DNA is made, fluorescent labelled and allowed to hybridize to DNA in the spots. Analysis of fluorescence made to draw conclusions.

Detailed Analysis

We consider the examples of the healthy and the infected apple. RNA is isolated from healthy apple, complementary DNA made and labelled with red fluorescent dye. RNA is isolated from infected apple, complementary DNA made and labelled with green fluorescent dye. Complementary DNA from the 2 tissues are mixed and applied to the microarray. Amount of red and green dye on each spot is measured by an instrument. If any gene is expressed at equal level in both tissue meaning equal amt. of c DNA from healthy tissue (red) and infected tissue (green) have hybridized to the spot, then the spot is shown as yellow. The spots on microarray that correspond to gene that is expressed in diseased apple will give green fluorescence. For a gene expressed only in healthy apple the spot will give red fluorescence. Thus, various gene expression patterns can be studied simultaneously.

Applications

study of gene expression patterns in:

·         different cell types 

-e.g. skin cell vs. brain cell

·         cell in different developmental stages

-e.g. embryonic skin cell vs. adult skin cell

·         normal cell vrs diseased cell

-e.g. embryonic skin cell vs. adult skin cell

·         environment-specific

-e.g. skin cell untreated vs. treated

·         drugs, toxins

Limitations

1.       Relatively new technology (10 years old).

2.       Still has technical problems (background).

3.       Poor reproducibility between investigators.

4.       Still mostly manual procedure.

5.       Relatively expensive.

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About Payal Ghoshal Banerjee

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