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Decoding the DNA j0283803.gif (5750 bytes)

Sequencing:

A sequencing method, developed by Sanger, uses dideoxynucleotides with the DNA sample and labeled phosphorus or sulfur.

The first step is:

	Separation of the two strands of DNA.
	Reanealing the DNA with a primer that complements a piece within the DNA.

Note: dNTP= deoxynucleotide triphosphate. There are 4 nucleotides that are made up of four bases: Adenine (A), Thymine (T), Cytosine (C) and Guanine (G). A always pairs with T and G always pairs with C. In RNA molecule, Thymine is substituted for Uracil (U) which still pairs with A.

	Extension of this primer by the enzyme DNA polymerase and deoxynucleotides (dNTP).
	Termination reaction using different nucleotide for each reaction (ddNTP). [Four separate incubations, where each sample containing the same mix, but different ddNTP]. Since the ddNTP do not have the 3'hydroxyl group, it cannot form a bond with the next phosphodiester group. Thus, the reaction is terminated.
	Fragments of the DNA are separated on a polyacrylamide gel electrophoresis.

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