Mutagenesis

Mutagenesis is a process by which the genetic information of an organism is changed in a stable manner, either in nature or experimentally by the use of chemicals or radiation. In nature mutagenesis may lead to cancer, but is also the force of Mutagenesis as a science was developed based on work done by Hermann Muller, Charlotte Auerbach and J. M. Robson in the first half of the 20th century.

Background
DNA may be modified, either naturally or artificially, by a number of physical, chemical and biological agents. Radiation and chemical mutagens were shown to be able to induce changes to the DNA. In the 1920s, X-ray was shown to cause mutations in fruit flies in work done by Hermann Muller. The mutations were also accompanied by observable changes in the chromosomes, and Muller further proposed mutation as the cause of cancer. In 1940s, Charlotte Auerbach and J. M. Robson, found that mustard gas can also cause mutations in fruit flies. In nature, mutagenesis is the mechanism whereby the mutation in the genome is acquired. The mutation may be beneficial or deleterious - it is the driving force of evolution, an organism may acquire new traits through genetic mutation, but mutation may also result in impaired function of the genes, and in severe cases, causing the death of the organism. In laboratory, however, mutagenesis is a useful technique for generating mutations that allow the functions of genes and gene products to be examine in detail, as well as producing proteins with improved characteristics or novel function. Initially the ability of radiation and chemical mutagens to cause mutation was exploited to generate random mutations, later techniques were developed to introduce specific mutations.

Experimental approaches in mutagenesis
Early approaches to mutagenesis rely on methods which are entirely random in the mutations produced. Cells may be exposed to UV radiation or mutagenic chemicals, and mutants with desired characteristic are then selected. For example, Escherichia coli may be exposed to UV radition, then plated onto agar medium. The colonies formed are then replica-plated, one in rich medium, another in minimal medium, and mutants that have specific nutritional requirement can then be identified by its inability to grow in minimal medium and isolated. A number of methods for generating random mutagenesis in specific protein were later developed to screen for mutants with interesting or improved properties. This may be done by using doped nucleotides in oligonucleotides synthesis, conducting a PCR reaction in conditions that enhance misincorporation of nucleotide thereby generating mutants, and more recently in techniques such as phage display. It is however desirable that specific changes can be introduced to the DNA. Analogs of nucleotides and other chemicals were first used to generate localized point mutations. Such chemicals may be aminopurine which induces AT to CG transition, while nitrosoguanidine,, bisulfite, , and N4-hydroxycytidine may induce GC to AT transition. These technique allows specific mutations to be engineered into a protein. Current techniques for site-specific mutation are commonly done by using mutagenic oligonucleotides in a primer extension reaction with DNA polymerase.

The directed approach may be done systematically in such technique as alanine scanning mutagenesis whereby residues are systematically mutated to alanine in order to identify residues important to the structure or function of a protein.

Types of mutagenesis

 * Directed mutagenesis
 * Insertional mutagenesis
 * PCR mutagenesis
 * Signature tagged mutagenesis
 * Site-directed mutagenesis
 * Transposon mutagenesis