Plasmids

Plasmids are small double-stranded DNA molecules, usually circular, that can exist independently of host chromosomes and are present in many bacteria (they are also present in some yeasts and other fungi). They have their own replication origins and are autonomously replicating and stably inherited.

• A replicon is a sequence in the DNA that has a replication origin and is capable of being replicated. Plasmids and bacterial chromosomes are separate replicons.
• Plasmids have relatively few genes, generally less than 30. Their genetic information is not essential to the host, and bacteria that lack them usually function normally.
• Single-copy plasmids produce only one copy per host cell. Multi copy plasmids may be present at concentrations of 40 or more per cell.
• Characteristically, plasmids can be eliminated from host cells in a process known as curing. Curing may occur spontaneously or be induced by treatments that inhibit plasmid replication while not affecting host cell reproduction.

Plasmids are classified based on their function in the host cell. Some of the important plasmids found in bacteria are:

1. F plasmid: A plasmid called the fertility or F factor plays a major role in conjugation in E. coli and was the first to be described. The F factor is about 100 kilobases long and bears genes responsible for cell attachment and plasmid transfer between specific bacterial strains during conjugation. (Conjugation refers the genetic recombination between two organisms through physical contact).
2. R Plasmid: Plasmids often confer antibiotic resistance on the bacteria that contain them. R factors or plasmids typically have genes that code for enzymes capable of destroying or modifying antibiotics. They are not usually integrated into the host chromosome. Genes coding for resistance to antibiotics such as amphicillin, chloramphenicol and kanamycin have been found in plasmids. Some R plasmids have only a single resistance gene, whereas others can have as many as eight.
3. Col plasmid: Bacteria also harbor plasmids with genes that may give them a competitive advantage in the microbial world. Bacteriocins are bacterial proteins that destroy other bacteria. They usually act only against closely related strains. Bacteriocins often kill cells by forming channels in the plasma membrane, thus increasing its permeability. They also may degrade DNA and RNA or attack peptidoglycan and weaken the cell wall. Col plasmids contain genes for the synthesis of bacteriocins known as colicins, which are directed against E. coli. Similar plasmids carry genes for bacteriocins against other species. For example, Col plasmids produce cloacins that kill Enterobacter species. Clearly the host is unaffected by the bacteriocin it produces.
4. Metabolic plasmids or degradative plasmids: They carry genes for enzymes that degrade substances such as aromaticcompounds (toluene), pesticides (2,4-dichlorophenoxyaceticacid), and sugars (lactose).
5. Virulence plasmids: The plasmids that make their hosts more pathogenic because the bacterium is better able to resist host defense or to produce toxins. For example, enterotoxigenic strains of E. coli cause traveler’s diarrhea because of a plasmid that codes for an enterotoxin.

Apart from these, some special plasmids are found in soil bacteria, which involve the plant-microbe interactions.

1. Sym plasmids: Symbiotic plasmids found in rhizobia, responsible for effective nodulation of this bacterium with host legume. The recognition, entry, nodule development and nitrogen fixation are mediated by this plasmid. As the size of some of the sym plasmids are 1.5 megabasepairs (nearly half of the size of bacterial chromosome) sometimes, they were referred as megaplasmids. The rhizobial cell may have 1-5 copies of sym plasmids.
2. Ti plasmids: Tumor-inducing (Ti) plasmids found in the soil bacterium, Agrobacteriumtumifaceans (close relative of Rhizobia) are responsible for tumor formation in various plant roots. Its size is about 200 kilobase pairs and one copy is found in the bacterial cells.

Eukaryotic DNA

Most of the DNA in eukaryotic cells is located in the nucleus, extensively folded into the familiar structures we know as chromosomes. Each chromosome contains a single linear DNA molecule associated with certain proteins. In prokaryotic cells, most or all of the genetic information resides in a single circular DNA molecule about a millimeter in length; this molecule lies, folded back on itself many times, in the central region of the cell. The genome of an organism comprises its entire complement of DNA.

A chromosome is an organized structure of DNA and protein found in cells. It is a single piece of coiled DNA containing many genes, regulatory elements and other nucleotide sequences. Chromosomes also contain DNA-bound proteins, which serve to package the DNA and control its functions.

Chromosomes vary widely between different organisms. Cells may contain more than one type of chromosome; for example, mitochondria in most eukaryotes and chloroplasts in plants have their own small chromosomes.

In eukaryotes, nuclear chromosomes are packaged by proteins into a condensed structure called chromatin. This allows the very long DNA molecules to fit into the cell nucleus. The structure of chromosomes and chromatin varies through the cell cycle. Chromosomes are the essential unit for cellular division and must be replicated, divided, and passed successfully to their daughter cells so as to ensure the genetic diversity and survival of their progeny. Chromosomes may exist as either duplicated or unduplicated. Unduplicated chromosomes are single linear strands, whereas duplicated chromosomes contain two identical copies (called chromatids) joined by a centromere.

Yeast genome – A model eukaryotic cell for genetics

Ascomycetous, non-filamentous (single celled) fungi and its metabolic and genetic features make it convenient model organism for studying the eukaryotic genetics. The features are

• Just 3.5 times bigger genome than E. coli (about 12 mb).
• Like bacteria, rapid growth and dispersed cells in the growth media.
• Well-defined genome organization.
• Mutation and screening mutants easy
• 16 chromosomes as haploid with a size of 200 to 2200 kb each.
• If needed, plasmids can be introduced like prokaryotic system.
• Controlled metabolisms (both oxidative and fermentative energy production)
• Have haploid (n=16) and diploid (2n = 32) stages
• Both vegetative (budding) and sexual reproduction (somatogamy)

Yeast genome

16 well-defined chromosomes with 200 to 2200 kb sized each with a total of 12,052 kbp.

There are 6183 genes (occupying 70%) are present.

Mitochrondrial DNA (as extrachromosomal) present.

Some yeast harbor plasmids (known as 2µm circle plasmids).

Some yeast harbor double strand RNA virus (which is responsible for killer toxin production).

Sexual reproduction and genetic variability

There are two types yeasts are present with reference to their sexual mode of reproduction.

• Homothallic yeast – Both male and female gametes (sex cells) are present in same thallus (body). Hence the fungus is self-fertile.
• Heterothallic yeast – Male gamete and female gamete are in different thallus and hence they are self-sterile.

S. cerevisiae exists in 3 stable developmental forms namely., Haploid ( 2 mating types called a & α) and diploids

MAT­a - haploid mating type a; MATα - haploid mating type α.

The homothallic fungus can switch their mating type from a to α and α to a and hence, self-fertilization is possible and form diploids even a single pure culture is maintained.

Whereas, the heterothallic yeast always maintains stable mating type and cannot form diploids when they are in single pure cultures.