Lecture notes on DNA -Microarrays

Microarrays are linear arrays of molecules immobilized at definite locations on an inert solid support such as a glass slide allowing them to be studied simultaneously. 

DNA microarrays and protein microarrays are widely being used in genomics and proteomics, respectively.

Principle:

Microarray technology is based on the principle of hybridisation between a 'probe' and target molecules that are to be detected. 

This is based on complementarity between the two molecules -such as the base pairing of complementary strands of DNA or protein-antibody interaction. 

The probes are attached to a solid support and the experimental sample is in solution.

 Once the target molecules bind the probes, these are detected by appropriate methods.

DNA Microarray

An array is an orderly arrangement of samples. 

It provides a medium for matching known and unknown DNA samples based on base-pairing. 

An array experiment can be conducted on substrates such as microplates or standard blotting membranes. 

To create an array, the sample may be deposited by hand or by robotics. 

In general, arrays are described as macroarrays or microarrays, the difference being the size of the sample spots. 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.

DNA microarrays or DNA chips, which have DNA molecules (the probes)

immobilized at precise locations on a substrate have become well accepted platforms for studying gene expression and tracking mutations.

 The solid substrate can be a silicon

wafer, a thin sheet of glass, plastic or a nylon membrane . 

Microarrays are prepared by arraying the samples in the cells of the substrate by high-speed robotics. 

In each reaction cell, trillions of polymeric molecules from a specific sequence of single strand DNA fragment are immobilized . 

These known sequences of single

strand DNA fragments immobilized on the substrates are often called probes. Unknown fragments of single stranded DNA samples, called the target, are allowed to react or hybridize with the probes on the chip.

 The double stranded DNA hybrids are formed in the chips where the target and the probe are complementary according to the base-pairing rule.

 To facilitate the diagnosis or analysis of the hybridized chip, the target samples are

often labelled with tags, such as fluorescent dyes (say, red and green), or radioisotope molecules.

 Spots with fluorescence can be detected using scanners interfaced with a

computer that generates an image of the array. 

The image is then processed using

appropriate software packages to generate microarray data.

Applications

DNA microarrays can be used to detect DNA sequences in samples.

 Microarray

technique can be used to detect cellular RNA indirectly (after reverse transcription

to DNA). 

This method of measuring gene expression via CDNA is called expression

analysis. 

Microarray technology has important applications in pharmacogenomics.

These include drug discovery and development, drug safety and molecular diagnostics.

DNA chips would also help evolve personalized medicines.