Aglo Transformation Lab Report
Introduction
When a bacterium integrates a piece of DNA into its genome, bacterial transformation has occurred. In this experiment bacterial transformation will be done using calcium chloride/heat shock. This is done by incorporating the plasmids into chemically competent cells that were made permeable by the calcium chloride solution and heat shock. In 1928, Frederick Griffith, a physician from London, was he first person to experiment with bacterial transformation. He permanently transformed a safe, nonpathogenic bacterial strain of pneumococcus into a deadly pathogenic strain. [1]
Plasmids are small circular autonomously replicating pieces of DNA that can be found inside of a prokaryotic bacterial cell. By barrowing a cell’s polymerase they replicate their own DNA. They are easy to extract from the bacterial cells due to their size. Plasmids are helpful for cloning foreign genes because of their ability to express antibiotic resistance as well their ability to be modified to express proteins of interest. A pGLO plasmid contains genes for the green florescent protein (GFP) as well as the gene for ampicillin resistance known as beta-lactamase. It also contains a gene regulation system (operon) that has the ability to control expression of the GFP gene in transformed cells known as araC. The source of GFP is naturally founds within a …show more content…
species of jellyfish, Aequorea Victoria. When exposed to UV light GFP will glow bright green due to light caused resonation. It is used as a marker or indicator in biotechnology, and has been used to create luminescent animals and plants. Its production is monitored by the pGLO plasmid’s arabinose operon. Within this operon the araC prevents the RNA polymerase from binding with the promoter, but with the arabinose it will instead promote the RNA polymerase. Once attached to the promoter the transcription of GFP will begin and stop when there is no more arabinose left. As for the ampicillin, it is an antibiotic that is a part of the penicillin drug group. It is used to fight various types of infections within the human body. It can be used for infections ranging from ear infections, to STD’s, to E.coli or salmonella making it particularly ideal for this experiment.
Escherichia coli, better known as E.
coli is a gram negative, bacterium found in the lower intestine of warm blooded organisms. Although E.coli strains are harmless there are serotypes that can cause food poisoning in humans. [2] Harmless strains can be found in the flora of the gut and produce vitamin k¬¬2 and prevent pathogenic bacteria from establishing and are therefore beneficial to the host. The major cause for disease is fecal to oral transmission. It is an ideal candidate for bacterial transformation because it is made of only one cell, reproduces every 20 minutes, is not harmful to people, and cannot survive outside the
lab.
When a bacterium integrates a piece of DNA into its genome, bacterial transformation has occurred. In this experiment bacterial transformation will be done using calcium chloride/heat shock. This is done by incorporating the plasmids into chemically competent cells that were made permeable by the calcium chloride solution and heat shock. In 1928, Frederick Griffith, a physician from London, was he first person to experiment with bacterial transformation. He permanently transformed a safe, nonpathogenic bacterial strain of pneumococcus into a deadly pathogenic strain. [1]
Plasmids are small circular autonomously replicating pieces of DNA that can be found inside of a prokaryotic bacterial cell. By barrowing a cell’s polymerase they replicate their own DNA. They are easy to extract from the bacterial cells due to their size. Plasmids are helpful for cloning foreign genes because of their ability to express antibiotic resistance as well their ability to be modified to express proteins of interest. A pGLO plasmid contains genes for the green florescent protein (GFP) as well as the gene for ampicillin resistance known as beta-lactamase. It also contains a gene regulation system (operon) that has the ability to control expression of the GFP gene in transformed cells known as araC. The source of GFP is naturally founds within a …show more content…
species of jellyfish, Aequorea Victoria. When exposed to UV light GFP will glow bright green due to light caused resonation. It is used as a marker or indicator in biotechnology, and has been used to create luminescent animals and plants. Its production is monitored by the pGLO plasmid’s arabinose operon. Within this operon the araC prevents the RNA polymerase from binding with the promoter, but with the arabinose it will instead promote the RNA polymerase. Once attached to the promoter the transcription of GFP will begin and stop when there is no more arabinose left. As for the ampicillin, it is an antibiotic that is a part of the penicillin drug group. It is used to fight various types of infections within the human body. It can be used for infections ranging from ear infections, to STD’s, to E.coli or salmonella making it particularly ideal for this experiment.
Escherichia coli, better known as E.
coli is a gram negative, bacterium found in the lower intestine of warm blooded organisms. Although E.coli strains are harmless there are serotypes that can cause food poisoning in humans. [2] Harmless strains can be found in the flora of the gut and produce vitamin k¬¬2 and prevent pathogenic bacteria from establishing and are therefore beneficial to the host. The major cause for disease is fecal to oral transmission. It is an ideal candidate for bacterial transformation because it is made of only one cell, reproduces every 20 minutes, is not harmful to people, and cannot survive outside the
lab.