DNA Sequence Analysis Methods

by Philip McIntosh

Overview

DNA Sequence Analysis Methods

Analysis of DNA sequences provide data on the genes, traits, identification and evolution of organisms. Initial sequencing techniques were slow, labor intensive and provided limited data. Current techniques rely on automation to sequence millions of bases in a matter of hours at low cost.
Chain Termination Sequencing

Chain termination uses chemically modified bases (A, T, C and G) in DNA synthesis reactions to generate DNA fragments that randomly terminate at one of the bases. The fragments are separated by size using gel electrophoresis, and the sequence is read directly off the gel (or a photo of the gel).

Maxam-Gilbert Sequencing

Maxam-Gilbert sequencing starts with full-length, radioactive or fluorescent labeled DNA. The DNA is cut into fragments using DNA cleaving reagents, then analyzed using electrophoresis.
Automated Sequencing

In automated DNA sequencing, robotic machines are often used to perform the reactions. The technique is usually a variation of the chain termination method using bases labeled with fluorescent dyes (a different color for each base). The final reaction products are separated using capillary electrophoresis, and the order of the bases is read optically as the fragments pass through a detector.
Shotgun Sequencing

The chain termination method is applicable to sequences of up to 1,000 bases. Shotgun sequencing is a method for generating sequences of entire genomes. Large DNA molecules are fragmented and sequenced using chain termination. The process is repeated to obtain a collection of short overlapping sequences, and the entire sequence is assembled using computer programs.
Direct Visualization

One of the more interesting (and speculative) methods on the horizon involves direct visualization of DNA using a combination of nanotechnology and electron microscopy techniques.

Overview