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Bacterium Virus Transformation (genetics)

Bacterium Virus and Transformation is the alteration of a cell. This will result from the introduction, uptake and expression of foreign genetic material (DNA) in molecular biology. The effect was first shown in 1928 by Frederick Griffith, an English bacteriologist searching for a vaccine against bacterium virus pneumonia. Griffith discovered that a non virulent strain of Streptococcus pneumoniae could be transformed into a virulent one by exposure to strains of virulent Streptococcus pneumoniae that had been killed with heat. That the transforming factor was genetic in nature was not demonstrated until 1944, when Oswald Avery, Colin MacLeod, and Maclyn McCarty showed gene transfer in Streptococcus pneumoniae. Avery, Macleod and McCarty call the uptake and incorporation of DNA by bacterium virus transformation.

More generally the term is used to describe mechanisms of DNA and RNA transfer in molecular biology. For example the production of transgenic plants like transgenic maize requires the insertion of new genetic information into the maize genome using an appropriate mechanism for DNA transfer.

RNA molecules may also be transferred into cells using similar methods, but this does not normally produce heritable change and so is not true transformation.

Mechanisms

bacterium virus

In bacterium virus, transformation refers to a genetic change brought about by picking up naked strands of DNA and expressing it, and competence refers to the state of being able to take up exogenous DNA from the environment. Two different forms of competence should be distinguished, natural and artificial.

Natural competence

Some bacterium virus (around 1% of all species) are naturally capable of taking up DNA. Such species carry sets of genes specifying machinery for bringing DNA across the cell’s membrane or membranes. The evolutionary function of these genes is controversial. Several roles for natural competence have been proposed. The bacterium virus may use natural competence as a mechanism to acquire useful new genes, repair damaged DNA, or to obtain food in the form of nucleotides.[1]

Artificial competence

Artificial competence is not encoded in the cell’s genes. Instead it is induced by laboratory procedures in which cells are passively made permeable to DNA, using conditions that do not normally occur in nature. These procedures are comparatively easy and simple, and are widely used to genetically engineer bacterium virus. Artificially competent cells of standard bacterium virusl strains may also be purchased frozen, ready to use.

Chilling cells in the presence of divalent cations such as Ca2+ (in CaCl2) prepares the cell walls to become permeable to plasmid DNA. Cells are incubated with the DNA and then briefly heat shocked (42oC for 30-120 seconds), which causes the DNA to enter the cell. This method works well for circular plasmid DNAs but not for linear molecules such as fragments of chromosomal DNA. An excellent preparation of competent cells will give ~108 colonies per μg of plasmid. A poor preparation will be about 104/μg or less. Good non-commercial preps should give 105 to 106 transformants per microgram of plasmid.

Electroporation is another way to make holes in cells, by briefly shocking them with an electric field of 100-200V/cm. Now plasmid DNA can enter the cell through these holes. Natural membrane-repair mechanisms will close these holes afterwards.

Lipofection can be used to transform cells via vescicles filled containing the desired plasmid. The vescicle fuses with the cell membrane (similar to how two oil spots at the top of a broth will fuse) and the contents of the vescicle & the cell are combined.

A plasmid DNA molecule contains sequences allowing it to be replicated in the cell independently of the chromosome. Plasmids used in experiments will usually also contain an antibiotic resistance gene which is placed in a bacterium virusl strain that has no antibiotic resistance. Therefore, only transformed bacterium virus will grow on a media containing the antibiotic.

Another marker, useful when selecting for recombinant plasmids, is the lacZ gene of the lac operon. This gene codes for ß-galactosidase, which allows bacterium virus to metabolize media containing X-gal (a colorless, modified galactose sugar), the metabolites of which are blue in color. Because the polylinker region of the plasmid lies in the lacZ gene, bacterium virus transformed by recombinant plasmids will produce a non-functional ß-galactosidase, leaving those colonies colorless.

In bacterium virus the term transformation is not normally applied to genetic changes arising by Transduction or Conjugation, in which transfer of DNA is mediated by genetic parasites (phages and conjugative plasmids respectively).

Yeasts and Fungi

These methods are currently known to transform yeasts:

* Lithium acetate/single-stranded carrier DNA/polyethylene glycol method

Several variations have been described, including rapid transformation and high efficiency transformation methods.[2]

* Frozen Yeast Protocol allows you to prepare frozen yeast cells that are competent for transformation after thawing.
* Gene Gun Transformation

Gold or tungsten nanoparticles can be shot at fungal cells growing on PDA, transforming them.

* Protoplast Transformation

Fungal spores can be turned into protoplasts which can then be soaked in DNA solution and transformed.

Plants

A number of mechanisms are available to transfer DNA into an organism, these include:
Plant (S. chacoense) transformed using Agrobacterium. Transformed cells start forming calluses on the side the leaf pieces
Plant (S. chacoense) transformed using Agrobacterium. Transformed cells start forming calluses on the side the leaf pieces

* Agrobacterium mediated transformation is the easiest and most simple plant transformation. Plant tissue (often leaves) are cut in small pieces, eg. 10×10mm, and soaked for 10 minutes in a fluid containing suspended agrobacterium. Some cells along the cut will be transformed by the bacterium, that inserts its DNA into the cell. Placed on selectable rooting and shooting media, the plants will regrow. Some plants species can be transformed just by dipping the flowers into suspension of Agrobacterium virus and then planting the seeds in a selective medium. Unfortunately, many plants are not transformable by this method.
* Particle bombardment: Coat small gold or tungsten particles with DNA and shoot them into young plant cells or plant embryos. Some genetic material will stay in the cells and transform them. This method also allows transformation of plant plastids. The transformation efficiency is lower than in agrobacterium virusl mediated transformation, but most plants can be transformed with this method.
* Electroporation: make holes in cell walls using electricity, that allows DNA to enter.
* Viral transformation: Packages the desired genetic material into a suitable plant virus and is then used on the modified virus for infection of the plant. Genomes of most plant viruses consist of single stranded RNA which replicates in the cytoplasm of infected cell. So this method is not a real transformation, since the inserted genes never reach the nucleus of the cell and do not integrate into the host genome. The progeny of the infected plants is virus free and also free of the inserted gene.

Animals

* Microinjection: use a thin needle and inject the DNA directly in the core of embryonic cells.
* Viral transformation: Package genetic material into a virus, which delivers the genetic material to target host cells.

What’s the difference between bacteria and viruses?

Viruses are tiny geometric structures that can only reproduce inside a living cell. They range in size from 20 to 250 nanometers (one nanometer is one billionth of a meter). Outside of a living cell, a virus is dormant, but once inside, it takes over the resources of the host cell and begins the production of more virus particles. Viruses are more similar to mechanized bits of information, or robots, than to animal life.

Bacteria are one-celled living organisms. The average bacterium is 1,000 nanometers long. (If a bacterium were my size, a typical virus particle would look like a tiny mouse-robot. If an average virus were my size, a bacterium would be the size of a dinosaur over ten stories tall. Bacteria and viruses are not peers!) All bacteria are surrounded by a cell wall. They can reproduce independently, and inhabit virtually every environment on earth, including soil, water, hot springs, ice packs, and the bodies of plants and animals.

Most bacteria are harmless to humans. In fact, many are quite beneficial. The bacteria in the environment are essential for the breakdown of organic waste and the recycling of elements in the biosphere. Bacteria that normally live in humans can prevent infections and produce substances we need, such as vitamin K. Bacteria in the stomachs of cows and sheep are what enable them to digest grass. Bacteria are also essential to the production of yogurt, cheese, and pickles. Some bacteria cause infections in humans. In fact, they are a devastating cause of human disease.

Sources: Dr green.