Phage genetics

Bacteriophages, viruses that infect bacteria , are useful in the study of how genes function. The attributes of bacteriophages include their small size and simplicity of genetic organization.

The most intensively studied bacteriophage is the phage called lambda. It is an important model system for the latent infection of mammalian cells by retroviruses , and it has been widely used for cloning purposes. Lambda is the prototype of a group of phages that are able to infect a cell and redirect the cell to become a factory for the production of new virus particles. This process ultimately results in the destruction of the host cell (lysis). This process is called the lytic cycle. On the other hand, lambda can infect a cell, direct the integration of its genome into the DNA of the host, and then reside there. Each time the host genome replicates, the viral genome undergoes replication, until such time as it activates and produces new virus particles and lysis occurs. This process is called the lysogenic cycle.

Lambda and other phages, which can establish lytic or lysogenic cycles, are called temperate phages. Other examples of temperate phages are bacteriophage mu and P1. Mu inserts randomly into the host chromosome causing insertional mutations where intergrations take place. The P1 genome exists in the host cell as an autonomous, self-replicating plasmid.

Phage gene expression during the lytic and lysogenic cycles uses the host RNA polymerase, as do other viruses. However, lambda is unique in using a type of regulation called antitermination.

As host RNA polymerase transcribes the lambda genome, two proteins are produced. They are called cro (for "control of repressor and other things") and N. If the lytic pathway is followed, transcription of the remainder of the viral genes occurs, and assembly of the virus particles will occur. The N protein functions in this process, ensuring that transcription does not terminate.

The path to lysogeny occurs differently, involving a protein called cI. The protein is a repressor and its function is to bind to operator sequences and prevent transcription. Expression of cI will induce the phage genome to integrate into the host genome. When integrated, only the cI will be produced, so as to maintain the lysogenic state.

The virus adopts the lytic or lysogenic path early following infection of the host bacterium. The fate of the viral genetic material is governed by a competition between the cro and cI proteins. Both can bind to the same operator region. The region has three binding zones—cro and cI occupy these zones in reverse order. The protein, which is able to occupy the preferred regions of the operator first, stimulates its further synthesis and blocks synthesis of the other protein.

Analysis of the genetics of phage activity is routinely accomplished using a plaque assay. When a phage infects a lawn or layer of bacterial cells growing on a flat surface, a clear zone of lysis can occur. The clear area is called a plaque.

Aside from their utility in the study of gene expression, phage genetics has been put to practical use as well. Cloning of the human insulin gene in bacteria was accomplished using a bacteriophage as a vector. The phage delivered to the bacterium a recombinant plasmid containing the insulin gene. M13, a single-stranded filamentous DNA bacteriophage, has long been used as a cloning vehicle for molecular biology . It is also valuable for use in DNA sequencing, because the viral particle contains single-stranded DNA, which is an ideal template for sequencing. T7 phage, which infects Escherichia coli, and some strains of Shigella and Pasteurella, is a popular vehicle for cloning of complimentary DNA. Also, the T7 promoter and RNA polymerase are in widespread use as a system for regulatable or high-level gene expression.

See also Bacteriophage and bacteriophage typing; Microbial genetics; Viral genetics