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DNA Transformation

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Melanie Grondin

on 25 February 2013

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Transcript of DNA Transformation

DNA Transformation Uses Bacteria History The process of single cells taking up foreign DNA from their environment and incorporating it into their own genetic codes. -host site for DNA cloning; allows for multiple copies of DNA to be created
-genetically modify bacteria Plants and Animals -determine the function significance of genes/amino acids
-genetically engineer plants/animals to add/remove certain proteins
e.g. insulin, which can be used to treat Type 1 Diabetes Avery et al. were the ones who discovered that the "transforming principle" was genetic material. They isolated DNA from the virulent strain and were able to transform harmless strains to virulent ones using just the DNA. How It Happens Bibliography http://www.biotechlearn.org.nz/themes/dna_lab/bacterial_transformation

http://www.clfs.umd.edu/classroom/bsci415/straney/lab/transformation.html

http://agwest.sk.ca/publications/infosource/inf_oct01.pdf

http://labtutorials.org/2011/03/16/transformation-of-competent-cells/

http://www.inia.es/gcontrec/pub/316-330(47_05-Methodologie_1165317453984.pdf

http://www.learner.org/courses/biology/textbook/gmo/gmo_5.html

http://en.wikipedia.org/wiki/Transformation_(genetics)

http://en.wikipedia.org/wiki/Natural_competence

http://www.dnalc.org/resources/animations/transformation2.html

http://www.annualreviews.org/doi/abs/10.1146/annurev-med-062310-094219?journalCode=med

revistas.um.es/hh/article/download/128191/119431

http://plato.acadiau.ca/courses/biol/Microbiology/transfer.htm#transformation

http://www.dnalc.org/resources/animations/transformation1.html

Raven and Johnson: Biology 7th Edition Types of Transformation - Natural Transformation: - Artificial Transformation: In Bacteria occurs in cells naturally, particularly bacteria In Eukaryotes occurs in laboratory; scientists introduce foreign
DNA into cells (typically E. coli) Transformation does not occur in many eukaryotic organisms because many are multicellular or are far too complex to ensure that every cell has taken up the same, intended genetic material. For transformation to happen, bacteria have to be in a certain physiological state called competence (eg. a time-limited response to environmental conditions such as starvation or DNA damage) Natural Competence Artificial Competence Most cells are generally unable to take up genetic information on their own. However some, such as B. subtilis, let foreign DNA through their membranes without the help of lab techniques. Most cells are not naturally competent and, instead, must be encouraged to accept artificial DNA through laboratory techniques. Electroporation and using Calcium Chloride are the two most common methods. Electroporation Calcium Chloride Bacteria take up DNA for the following reasons:
1. Genetic Diversity
- new virulence traits, or antibiotic resistance
2. DNA Repair
- DNA from closely related species can serve as template for DNA repair
3. Food
- source of carbon, nitrogen and phosphorous Therefore, the need for nucleotides to synthesize RNA and DNA can cause a bacterial cell to become naturally competent.
Also, DNA damaging conditions induces competence in a cell. For example, exposure to UV light, a DNA damaging agent. By electrifying the plasma membrane of a cell, you can alter its permeability by creating hydrophilic pores. How does this work? DNA is a polar molecule and, like water, does not easily pass through the hydrophobic regions of the membrane. However, once pores are made within the membrane, the genetic material can easily pass through. The more coiled the DNA is, the better its chances of being taken up during transformation are. This is why plasmids are used instead of linear DNA fragments. Also the DNA usually has an antibiotic-resistance marker, to help identify the successfully transformed bacteria. (chemical competence) It can still work! Transformation is still used to alter plant cells. Many plants can regrow from a single cell, making them optimal for genetic modification via transformation. Transformation can also occur naturally in parts of plants, though it doesn't usually affect the whole plant. DNA cannot pass through the cell membrane because the negatively-charged phosphates on the DNA helix are repelled by those on the lipids of the membrane. Lab Methodology Gene Gun The calcium ions from calcium chloride interacts with the negative charges and creates an electrostatically neutral situation. Lowering the temperature solidifies the lipids in the membrane and stabilizes the negatively-charged phosphates A rapid rise in temperature (heat shock) creates a temperature imbalance on both sides of the membrane causing a current. This allows the DNA to go through the cell. DNA transformation was first observed by Griffith in 1928, when he discovered that harmless Streptococcus pneumonia can turn virulent if exposed to heat-killed virulent strains. He hypothesized that there was a "transforming principle" in the virulent strain that is causing the change in non-virulent strains. Frederick Griffith Experiment Avery-MacLeod-McCarty Experiment Small beads made of tungsten or gold are covered with DNA and shot into the desired cell. Electroporation Microinjection A very fine needle is used to inject the desired cell with genes coding for desired traits. In Nature Agrobacterium tumefaciens and related bacteria are often sited to cause DNA transformation within plants. They are found in soil and will aggregate around injured areas of the plant. This particular strain of bacteria causes tumors to form on plants. This bacteria is often manipulated in order to introduce helpful DNA into plant cells. In Animals Any DNA transformation done on animals is usually called transfection. Cancer and Metastasis Metastasis is the spreading of a primary cancer to a secondary location. For example, lung cancer may metastasize to the liver, causing lung cancer tumors to form there. Though it is generally accepted that cancer metastasizes through the movement of entire tumor cells, there has been some studies that show that tumor DNA in the bloodstream can also cause cancer to spread via DNA transformation. However, this phenomenon is relatively under-researched. Process of Transformation After the bacteria cells lyses, they release their DNA as fragments into the environment.
DNA can also be placed in the environment by scientists.
Living bacteria can absorb these fragments and incorporate them into their DNA by recombination. The fragments that are not absorbed degrade through time.

This method of transfer is the process of transformation. Recombinant Cell The genetically transformed cell is called a recombinant cell because it has a different genetic makeup than the donor and the recipient. Please Note: The makers of this Prezi: Dalia, Melanie, Weiwei, and Wanchen, are fully aware that species names should be italicized. However, despite our great technological prowess, we could not figure out how to do that on Prezi.
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