Transformation Of Escherichia Coli With pGLO Plasmid
Transformation Of Escherichia Coli With pGLO Plasmid
April 24, 2013
ABSTRACT: This experiment focuses on genetic engineering and transformation of bacteria. The characteristics of bacteria are altered from an external source to allow them to express a new trait, in this case antibiotic resistance. In is experiment foreign DNA is inserted into Escherichia coli in order to alter its phenotype. The goal of the experiment is to transform E. coli with pGLO plasmid, which carries a gene for ampicillin resistance, and determine the transformation efficiency. The bacteria are transformed by a combination of calcium chloride and heat shock. When the bacteria are incubated on ice, the fluid cell membrane is slowed and then the heat shock increases permeability of the membrane. The results obtained in the experiment show that the E. coli that was transformed with pGLO was able to resist ampicillin and grow in its presence. These results suggest that microorganisms can be genetically engineered to selectively resist certain contaminants, which means that they can potentially be used to human benefit to rid the environment, or even the human body, of unwanted toxins.
INTRODUCTION:
Transformation occurs when altered genetic characteristics of bacteria are acquired from a different source. Plasmids are used to transform bacteria because they are small pieces of DNA capable of independently replicating and therefore transferring their (often beneficial) traits to the bacteria. The goal of genetic transformation in this experiment is for the bacteria Escherichia coli to obtain an antibiotic resistance to ampicillin, which can be physically observed when the bacteria expresses the reporter gene Green Fluorescent Protein (GFP) because the transformed bacteria will glow green under UV light when in the presence of arabinose. The gene for GFP is naturally found in a bioluminescent jellyfish, allowing it to glow in the dark. The plasmid used to transform the bacteria
April 24, 2013
ABSTRACT: This experiment focuses on genetic engineering and transformation of bacteria. The characteristics of bacteria are altered from an external source to allow them to express a new trait, in this case antibiotic resistance. In is experiment foreign DNA is inserted into Escherichia coli in order to alter its phenotype. The goal of the experiment is to transform E. coli with pGLO plasmid, which carries a gene for ampicillin resistance, and determine the transformation efficiency. The bacteria are transformed by a combination of calcium chloride and heat shock. When the bacteria are incubated on ice, the fluid cell membrane is slowed and then the heat shock increases permeability of the membrane. The results obtained in the experiment show that the E. coli that was transformed with pGLO was able to resist ampicillin and grow in its presence. These results suggest that microorganisms can be genetically engineered to selectively resist certain contaminants, which means that they can potentially be used to human benefit to rid the environment, or even the human body, of unwanted toxins.
INTRODUCTION:
Transformation occurs when altered genetic characteristics of bacteria are acquired from a different source. Plasmids are used to transform bacteria because they are small pieces of DNA capable of independently replicating and therefore transferring their (often beneficial) traits to the bacteria. The goal of genetic transformation in this experiment is for the bacteria Escherichia coli to obtain an antibiotic resistance to ampicillin, which can be physically observed when the bacteria expresses the reporter gene Green Fluorescent Protein (GFP) because the transformed bacteria will glow green under UV light when in the presence of arabinose. The gene for GFP is naturally found in a bioluminescent jellyfish, allowing it to glow in the dark. The plasmid used to transform the bacteria
Cited: Chung CT. 1989. One-step preparation of competent Escherichia coli: Transformation and storage of bacterial cells in the same solution. Proceedings of the National Academy of Sciences of the United States of America, 86:2172-2175. Hanahan D. 1983. Studies on Transformation of Escherichia coli with Plasmids. Journal of Molecular Biology, 166:557-580. Pieper DH. 2000. Engineering bacteria for bioremediation. Current Opinion in Biotechnology, 11.3:262-270. Spilios K (ed). 2013. Principles of Biology, II. Hayden-McNeil Publishing, Plymouth, MI. Module #6, pp 119-127.