Prokaryotic Genetics Tools

What biological phenomena do we use as tools to study prokaryotic genetics?

Bacterial Conjugation

Bacteria , in general, reproduce asexually, but in order to increase diversity and share the gene pool, they have developed a mechanism for transfer of genetic material from one bacterium to another.

The ability to perform this transfer is conferred by a set of genes which are called F for fertility. These genes can exist on a small, circular piece of DNA that replicates independently from the bacterial chromosome, or they can be integrated into the chromosome. The bacterium containing this gene (often parochially called the "male" bacterium) shoots a grappling hook (called a pilus) out to sieze a neighbouring bacterium. The two cells then are drawn together, and the grappling hook forms a channel through whch DNA is transferred. This process is called bacterial conjugation.

Microbial biologists have learned to take advantage of this phenomenon. When you were studying mendelian genetics, you could often analyse the function of a gene by looking at how it related to its allele pair: is the allele you are looking at recessive or dominant? In a haploid organism like the prokaryote, this analysis is impossible. However, pseudo-diploid (meridiploid) genetics can be conducted by putting various alleles of a gene of interest on the F element, along with the fertility genes. That way the genes can be transferred to another bacterium, which might already contain different alleles of those genes. The effect of the two alleles on each other can then be examined.

There are three manifestations of the fertility factor:

Manifestation Description
Hfr The F element has become integrated into the genome.
When conjugation occurs, the F genes start travelling across the pilus,
dragging the rest of the genome behind it. Eventually, the pilus breaks,
so most often the entire genome is not transferred. The bacterial genome can
be measured in minutes from the origin of transfer: The amount of time it takes
for a particular gene to be transferred from one bacterium
to another indicates how far it is from the origin of replication.
F' Also called the F' episome. This is a small circular piece of DNA
that contains the fertility genes and a few other genes. These other
genes are transferred very efficiently from one bacterium to the next
because the length of the transferred DNA is short enough that it can
move across the breach before the pilus breaks.
F This is a small circular piece of DNA carrying only the fertility genes.

Phage and Phage Transduction

There has already been a discussion of the phage cycle. Microbiologists have taken advantage of the lysogenic cycle of the phage to generate pseudo-diploid genetics in the haploid prokaryote. Particular genes can be packaged into phage heads and these phage can be used to infect bacteria. Phage strains have been constructed that cannot enter the lytic phase, so upon infection, the DNA integrates into the genome. If the bacterium already has a copy of that gene, the two alleles' effect on each other can be examined.

Transformation

Bacteria can be induced to take up small pieces of circular DNA from the medium by treating them with a solution of calcium chloride and then shocking them at 42° C. The mechanism of this absorption of DNA is not well understood, but if a piece of DNA is constructed by the methods of recombinant DNA manipulation to include particular genes of interest, the DNA can be transformed into bacteria cells containing another allele of the genes and the alleles' effect on each other can be examined.

Cross-feeding

If two bacteria strains that have different genotypes are plated near each other on the same petri dish, both will exude particular substances into the medium. If the product of the gene of interest can diffuse out of the cell, the product of one allele in one strain of bacteria can affect the product of a different allele in another strain.

hyperbio@mit.edu

Manifestation	Description
Hfr	The F element has become integrated into the genome. When conjugation occurs, the F genes start travelling across the pilus, dragging the rest of the genome behind it. Eventually, the pilus breaks, so most often the entire genome is not transferred. The bacterial genome can be measured in minutes from the origin of transfer: The amount of time it takes for a particular gene to be transferred from one bacterium to another indicates how far it is from the origin of replication.
F'	Also called the F' episome. This is a small circular piece of DNA that contains the fertility genes and a few other genes. These other genes are transferred very efficiently from one bacterium to the next because the length of the transferred DNA is short enough that it can move across the breach before the pilus breaks.
F	This is a small circular piece of DNA carrying only the fertility genes.