The ability to plate out large numbers of haploid bacteria or fungi on a Petri dish, and to examine a single colony (or clone) under a variety of conditions (with an without a growth factor, with and without a drug, or at high and low temperature), makes it relatively easy to screen through many individuals to find mutants with a particular phenotype. However, in order to carry out a complementation analysis, one needs to be able to combine the two haploid mutants in one cell. Many fungi, such as yeast, do this thorough a natural meiotic sporulation and mating process. Figure 1.6 illustrates the use of fungal matings in complementation.
Bacteria can also, although not by meiosis and fertilization, and only a part of the genome of one bacterium is transferred to another. The sexual transfer of information in E. coliuses plasmids called F (fertility) factors or Hfr strains. Male E. coli cells have a large plasmid, the F or fertility factor. A plasmidis a circular, extrachromosomal DNA molecule that is not essential to the bacterium. The F plasmid can transfer DNA from the male cell to an F- or female cell, in a process called conjugation (Figure 1.10). The male and female cells are brought close together by attachments at pili, the cells join and DNA is synthesized from the F plasmid and transferred into the recipient cells. This converts the female cell to a male cell, in response to conjugation via pili.
In some strains of E. coli the F factor is integrated. In this case, the DNA transfer starts in F region of the chromosome, but it also transfers adjacent chromosomal DNA. These are called hfr strains, for their high frequency of recombination. The transferred DNA recombines with the DNA in the recipient cell. Some F-related plasmids are a hybrid of F DNA and host bacterial DNA. These F’plasmids appear to be derived from F factors but they have replaced some of the F DNA with bacterial DNA. Thus they are convenient carriers of parts of the E. coli genome.
This conjugal transfer can be used to create partial diploids, also called merodiploids, in E. coli. For some time after conjugation, a portion of two different copies of the chromosome is present in the same cells. Another method is to introduce F’ factors, carrying bacterial DNA, into another strain. These are two ways to do complementation analysis in E. coli.
Figure 1.10. F-factor mediated conjugal transfer of DNA in bacteria.