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Modern Genetic Analysis Chapter 9. The Genetics of Bacteria and Phages
Transduction

Some phages are able to carry bacterial genes from one bacterial cell to another, a process called transduction ("leading across"). Phages are propagated on one bacterial genotype (the donor), and then progeny phage particles are collected. Most of these progeny are exact duplicates of the original infecting phage; however, a small fraction of them carry fragments of host DNA. The progeny phage population is used to infect another bacterial genotype (the recipient). Many of the phages that carry donor DNA fragments will infect a recipient. The phage DNA and any donor DNA are injected. Hence, in these cases, a temporary donorrecipient merozygote is created. Subsequently, the transduced bacterial genes may be incorporated into the bacterial chromosome by recombination (Figure 9-22). The presence of transduced donor alleles can be detected by plating on selective medium.

There are two kinds of transduction: generalized and specialized. Some phages can carry any part of the bacterial chromosome; such phages are generalized transducing phages. In contrast, specialized transducing phages carry only specific parts of the bacterial chromosome.

Phages P1 and P22 both show generalized transduction. These temperate phages have different fates in the cell: P22 inserts into the host chromosome as a prophage, whereas P1 remains free like a large plasmid. However, both act as generalized transducers because, in the course of lysis, bacterial DNA is accidentally "stuffed" into the phage head. Inside the recipient cell, the transducing fragments integrate by double crossover utilizing the host recombination system.

Phage l is a good example of a specialized transducer. We learned in the preceding section that l always inserts between the gal and bio genes in the E. coli host chromosome. The prophage normally excises by an outlooping mechanism that is a precise reversal of its mechanism of integration. It is able to transduce only when an occasional faulty outlooping includes the gal or bio genes into the phage circle. Let's trace the details of l transduction.

In Figure 9-23a, we see a schematic representation of the production of a lysogen, or lysogenic bacterium. The lytic cycle can be induced by ultraviolet light to produce a lysate (a population of progeny phages). The normal outlooping of the prophage restores the original phage chromosome (Figure 9-23b, i). These phages can integrate normally (as in Figure 9-23a) upon subsequent infection of a bacterial strain that is not lysogenic for l .

Rare outlooping can result in a phage particle that carries the gal gene (Figure 9-23b, ii). These particles are defective in that some phage genes have been left behind in the host; consequently, they are called l dgal or l -defective gal. The ldgal particle has a l protein coat and tail fibers and can infect bacteria, but it is defective in its integration site. The hybrid integration site left in ldgal does not afford a correct substrate for the phage-encoded enzyme that promotes recombination between the phage particle and bacterial genomes. Therefore, efficient integration cannot take place in a single infection of E. coli by ldgal. However, coinfection with a wild-type l phage results in efficient integration of ldgal (Figure 9-23c, i); the second, wild-type phage acts as a helper phage.

In practice, the lysate of l produced originally (Figure 9-23b) can be used to infect a gal recipient culture that is nonlysogenic for l . These infected cells are plated on a minimal medium, and rare gal+ transductants are the only ones to grow and produce colonies. A small percentage of these transductants result from recombination between the gal regions of the ldgal phage particle and the recipient chromosome (Figure 9-23c, ii). A majority of the transductants (Figure 9-23c, i) are double lysogens.

Specialized transduction can mobilize only small regions of the bacterial genome that flank the prophage. Transduction by l can mobilize either the gal or the bio gene, because gal and bio are the genes that can be incorporated into l by the rare, aberrant excision events. Specialized transduction is useful for moving specific genes from one bacterial strain to another.

Determining Linkage from Transduction

Generalized transduction can be used to derive linkage information about bacterial genes in cases in which genes are close enough that the phage can pick them up and transduce them as a single piece of DNA, an outcome known as cotransduction. For example, suppose we wanted to measure the linkage between met and arg in E. coli. An appropriate generalized transducing phage to use would be phage P1. Cotransduction by P1 can take place when genes are within approximately 1.5 minutes of each other on the E. coli chromosome map. We would allow phage P1 to infect a donor met+ arg+ strain, then collect P1 progeny, and use them to infect a recipient strain of genotype met arg. Then we would select for one donor genesay, met+by plating cells on medium that lacks methio-nine and selecting met+ colonies. These colonies would then be tested for the unselected marker to see what proportion of them is also arg+; in this way, we could determine the percentage of met+ arg+ cotransductants. Linkages are expressed as cotransduction frequencies (Figure 9-24). For example, if 20 percent of met+ colonies are found to be also arg+ (that is, 20 percent are met+ arg+ and 80 percent are met+ arg), then the cotransduction frequency is 20 percent. The met+ arg+ cotransducatants must have arisen from a transducing DNA fragment that contained both genes, both of which were inserted into the recipient chromosome by two crossovers flanking these loci. The met+ arg transductants may have arisen from the same type of fragment but with only the met+ gene inserted by crossovers flanking the met locus. It is also possible that the met+ arg category could have arisen from a transducing fragment that did not include arg+.

Note that, in this type of mapping, there is an inverse relation quite different from that obtained by using map units based on recombinant frequencies: the greater the cotransduction frequency, the closer two genetic markers are and the shorter is the map distance. For any given phage, cotransduction frequencies can be calibrated in terms of bacterial map units (minutes) by using empirical formulas.

MESSAGE
The frequency of cotransduction is inversely proportional to the map distance between the two genes concerned.

© 1999 by W. H. Freeman and Company. All rights reserved.