Diversity in Genetic Structure and Composition

The host cell's genetic material is destroyed.

Phage proteins

Phage nucleic acid enters the cell

Phage proteins are synthesized, nucleic acids are replicated, and host cell genetic material is destroyed.

Figure 5.1 Lytic cycle of bacteriophage

Phages can only multiply when grown within bacterial cells, thus depleting the growth medium of nutrients, limiting phage multiplication and spot size. Since each spot is the result of infection with a single initial phage particle, the number of individual spots present on the medium can be counted (Figure 5.2).

The genotypes of phage mutants can be determined by studying the spots. In some cases, the presence of spots is sufficient. For example, phage mutations that reduce the number of progeny phages from infected cells often produce smaller spots. Large spots may be produced by mutations that cause early lysis of infected cells, so that each spot continues to infect more rapidly. Another type of phage mutation is

can be determined by the ability or inability of phages to form spots on particular bacterial strains.

Figure 5.2 Phage entry into host cells in both parental forms simultaneously.

r + : small spot, r - : large spot, h + : faint spot, h - : clear spot

2. Genetic recombination in the lytic cycle

Phages are small in size and must be seen under an electron microscope. But the characteristics of phages are observed based on the spots or host amplitude. Let two phage T 4 strains with different genotypes infect an E. coli bacteria cell , some of the next generation phages will perform genetic recombination, phage DNA replicates and exchanges segments if there is negative interference. Allele r - dissolves quickly, resulting in large spots, allele h - infects host cells, resulting in clear spots. The cross is as follows:

r - h +  r + h -

Four types of spots are obtained. The two large, opaque spots and the small, clear spots correspond to the phenotypes of the parental phage. The other two phenotypes, large clear spots and small opaque spots, are recombinant forms corresponding to the genotypes r - h - and r + h + . When many bacteria are infected, a number of reversible recombinant forms are usually found among the phages in the next generation. In the experiment, each of the four genotypes produces a different phenotype in terms of spot type (Figure 5.2) . The number of genotypes can be determined by examining the spots formed. The recombinant frequency, expressed as a percentage, is determined as follows:

Recombination frequency =

3. Arrangement of genes in phage chromosomes

Recombination frequencies can be used to determine the distance of a map in eukaryotes. Mapping experiments showed that mutations in T4 mapped into three distinct clusters. All three clusters were linked to one another. George Streisinger et al. (1964) demonstrated that the genetic map of phage T4 was circular.

Membrane

Tail thread

Nucleotide exchange

DNA synthesis, replication and modification

Linkage group of genes determines origin

Tail root disc

Head, neck, neck folds

Tail root disc

Head

Figure 5.3 Genetic map of T4 with markers

In each cross, three to four genetic markers were set up in turn for each group and run through the entire T4 genome. Several other genes were identified and mapped completely on the circular molecule (Figure 5.3) . The regions in the inner circle are the three clusters of T4 markers that have been identified and mapped. The outer circle contains several large sets of markers that complete the entire circle of the genetic map. The T4 phage genetic map shows that the T4 phage genes cluster extensively according to their function. For example, there is a large cluster of genes for DNA replication in the fourth position.

on the upper right and has the gene cluster that synthesizes the components that make up the phage head at the bottom of the circle.

The T4 phage DNA molecule is a linear single-stranded molecule, with each end of the T4 phage DNA being either replicated or terminally redundant. Thus, each DNA molecule increases in size by 2%. As the DNA is replicated in the cell, recombination between the terminal portions of the T4 genome and homologous sequences of other T4 genomes results in a DNA product that is larger than the head can contain. The duplicated molecules are formed because recombination in the T4 phage genome occurs frequently, with an average of about 20% of recombination events occurring on a chromosome. When the DNA molecule is packaged into the head, it is enzymatically cleaved to contain only about 102% of the length of the T4 phage genome, because of the head repeat.

4. Mapping the fine structure of the rII region of phage T4

Each mutant can recombine with other mutants. Deletion mutations prevent recombination with two or more point mutations at different locations in the gene. Each deletion causes the loss of a portion of the phage genome including the rII region. Deletion mapping is a simple method for mapping thousands of mutants. Deletion mapping is based on the presence or absence of recombinant forms. In any cross between an unknown point mutant and a deletion mutant, the appearance of the wild-type form indicates that the point mutation is outside the deletion region. Conversely, if the point mutation occurs within the deletion region, no wild-type recombinant forms will appear in the next generation.

Several crosses were performed to map the detailed mutation of the rII gene. The distances from A1 to A6 and B are shown in Figure 5.4 . A particular mutation was examined located in the A4 region. This mutant did not recombine wild-type in crosses with large deletion mutants such as r1272, r1241, rJ3 and rPT1 but it could recombine wild-type in crosses with rPB242, rA105 and r638. Mutants generated by the same template, resulting from crosses with large deletion mutants, will be ranked