Whole Genome Sequencing

The techniques for gene transfer described in this chapter have allowed the construction of genetic maps for E. coli and a few other well investigated bacteria. However, for the vast majority of microorganisms, no "classical" genetics exists. Nowadays these are largely being investigated by more modern techniques, such as gene cloning and DNA sequencing.

Since the development of rapid automated techniques for sequencing DNA (see Ch. 24) many whole genomes have been totally sequenced. The first genome sequence to be finished was Hemophilus influenzae, in 1995. Since 1995, nearly 50 complete bacterial genomes have been sequenced, and another 100 are partially sequenced. Sequence comparison with genes of well-investigated organisms allows provisional identification of many genes. However, even in E. coli, the function of about a third of the genes remains uncertain.

Whole genome sequencing of pathogenic bacteria and comparison with their harmless relatives may reveal extra blocks of genes responsible for causing disease. Many virulence genes are carried on plasmids as discussed in Chapter 16. Others are found clustered together in regions of the chromosome known as "pathogenicity islands". Most genes of Salmonella, as well as their order around the chromosome, correspond to those of its close relative E. coli, as would be expected. However, extra segments of DNA are found in Salmonella that are lacking in E. coli. Some of these are pathogenicity islands (Fig. 18.22). Such extra regions are often flanked by inverted repeats, implying that the whole region was inserted into the chromosome by transposition at some period in the evolutionary past. In agreement with this idea, such islands are often found in some strains of a particular species but not others. In addition, these islands tend to have different GC to AT ratios and/or codon usage frequencies from the rest of the chromosome, suggesting their origin in some other organism. Conversely, E. coli possesses a few DNA segments missing in Salmonella. Interestingly, one of these is the area including the lac operon and a few surrounding genes. Thus the classic lac operon, the most-studied "typical" gene of the "standard organism" is probably a relatively recent intruder into the E. coli genome!

Pathogenicity islands are simply the best known case of "specialization islands". These are blocks of contiguous genes, presumed to have a "foreign" origin, that con-

pathogenicity island Region of bacterial chromosome containing clustered genes for virulence

The bacterium, Hemophilus influenzae, was the first organism to have its DNA completely sequenced.

Virulence genes are often clustered together forming "islands".

Differences in GC/AT ratios reveal segments of chromosomes with foreign origins.

FIGURE 18.22 Pathogenicity Islands of Salmonella

Comparison of the E. coli genome with its close relative, Salmonella, reveals large regions of DNA that have no homology (orange). The remaining regions have similar genes that are in identical order. For example, Salmonella genes d through j are clustered together in the exact same order as E. coli genes d through j. Since Salmonella is pathogenic and E. coli is not, the regions of no homology probably encode the genes required for pathogenicity; therefore, they are termed pathogenicity islands. The islands are flanked by inverted repeats, suggesting the DNA may have been acquired through transposition. Note: this figure is not drawn to scale; the pathogenicity islands are greatly exaggerated relative to the rest of the chromosome for purposes of illustration.

Bacterial chromosomes

Horizontal transfer of genes is especially significant in bacteria.

tribute to some specialized function that is not needed for simple survival. Not surprisingly, medical relevance has drawn most human interest. Other examples include genes encoding pathways for the biodegradation of aromatic hydrocarbons, herbicides and other products of human industry and pollution.

Movement of genes "sideways" is designated lateral or horizontal gene transfer in distinction to the "vertical" transfer of genes from ancestors to their direct descen-dents. Horizontal gene transfer can occur by natural transformation, viral transduc-tion, or transposon jumping. Horizontal gene transfer may occur between closely related organisms or those far apart taxonomically. Estimates suggest that in typical bacteria around 5% of the genes have been obtained by lateral gene transfer, and in rare cases up to 25%. Thermotoga is a eubacterium adapted to life at very high temperatures and which consequently shares its habitat with several archaebacteria. Ther-motoga has apparently gained around 25% of its genes by transfer from thermophilic archaebacteria such as Archaeoglobus and Pyrococcus. When we remember that the F-plasmid of E. coli can mediate DNA transfer into yeast (see Ch. 16), these results are perhaps not so surprising.

horizontal gene transfer Movement of genes sideways between unrelated organisms. Same as lateral gene transfer lateral gene transfer Movement of genes sideways between unrelated organisms. Same as horizontal gene transfer

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