RLO: DNA

• Introduction
• Hybridisation
• Fluorescent In Situ Hybridisation
• The Polymerase Chain Reaction
• DNA Sequencing
• DNA Microarrays

DNA Sequencing

Once a piece of DNA has been amplified, it can be sequenced. In this process, four reaction mixtures are set up, each one including:

1. DNA to be sequenced
2. DNA polymerase
3. A supply of nucleotides (A, C, G and T)
4. A small amount of a labelled chain-terminating variant of one of the four nucleotides.

The enzyme DNA polymerase incorporates a chain-terminating variant at random, eventually ending the chain at every possible nucleotide position over a few hundred bases.

The products of the reaction mixtures are run on an electrophoresis gel, where the sequence can be deduced by reading from the smallest to the largest piece.

If different fluorescent labels are used for the variant bases, sequencing can all be done in one single reaction, the bands can be detected and the sequence read out automatically. The entire process of DNA sample preparation and sequencing is now highly automated, a development that has been essential for the timely and cost-effective completion of the human genome project.