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Deoxyribonucleic acid (DNA) is the chemical compound that contains the instructions needed to develop and direct the activities of nearly all living organisms. DNA molecules are made of two twisting, paired strands, often referred to as a double helix.

Each DNA strand is made of four chemical units, called nucleotide bases, which comprise the genetic "alphabet." The bases are adenine (A), thymine (T), guanine (G), and cytosine (C). Bases on opposite strands pair specifically: an A always pairs with a T; a C always pairs with a G. The order of the As, Ts, Cs, and Gs determines the meaning of the information encoded in that part of the DNA molecule just as the order of letters determines the meaning of a word.

DNA Sequencing

Sequencing simply means determining the exact order of the bases in a strand of DNA. Because bases exist as pairs, and the identity of one of the bases in the pair determines the other member of the pair, researchers do not have to report both bases of the pair.

In the most common type of sequencing used today, called the chain termination method, a DNA strand is treated with a variety of nucleotides, a set of enzymes, and a specific primer to generate a collection of smaller DNA fragments. Four fluorescent tags, each specific for a given base, is part of the mixture. Each of the fragments differs in length by one base and is marked with a fluorescent tag that identifies the last base of the fragment. The fragments are then separated according to size and passed by a detector that reads the fluorescent tag. Then, a computer reconstructs the entire sequence of the long DNA strand by identifying the base at each position from the size of each fragment and the particular fluorescent signal at its end.

At present, this technology only can determine the order of up to 800 base pairs of DNA at a time. So, to assemble the sequence of all the bases in a large piece of DNA, such as a gene, researchers need to read the sequence of overlapping segments. This allows the longer sequence to be assembled from shorter pieces, somewhat like putting together a linear jigsaw puzzle. In this process, each base has to be read not just once, but at least several times in the overlapping segments to ensure accuracy.

Researchers can use DNA sequencing to search for genetic variations and/or mutations that may play a role in the development or progression of a disease. The disease-causing change may be as small as the substitution, deletion, or addition of a single base pair or as large as a deletion of thousands of bases.


Courtesy: National Human Genome Research Institute, United States of America accessed online at http://www.genome.gov/18016863 on September 16, 2011