6a lab

6a lab - Dhaval Rana Ap Biology December 16, 2009 Lab 6a:...

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Dhaval Rana Ap Biology December 16, 2009 Lab 6a: Transformation of E.coli with an Antibiotic Resistant Gene Abstract The objective of this lab is to demonstrate phenotypic changes in bacteria that have been transformed with an antibiotic- resistant gene and a metabolic marker. The hypothesis is that if the transformed E. coli contains pBLU plasmids, then blue colonies of the bacteria will be formed in the Petri dish. In this lab, we use plasmids that carry the ampR gene to transform E. coli cells that lack this gene. We also prepare a second group of E. coli cells as a control to verify that E. coli will not grow on agar with ampicillin unless it is transformed, and that nothing in the procedure itself affects the survival of E. coli. The procedure is that same for both groups of cells except pBLU plasmids is added to the experimental cells but not to the control cells. If there is no ampicillin in the agar, E. coli will cover the plate with so many cells it is called a “lawn” of cells. Only transformed cells can grow on agar with ampicillin. Since only some of the cells exposed to the pBLU plasmids will actually take them in, only some cells will be transformed. Thus there will be only individual colonies on the plate. If none of the sensitive E. coli cells have been transformed, nothing will grow on the agar with ampicillin. Introduction Genetic transformation occurs when a host organism takes in foreign DNA and expresses the foreign gene. In this part of the lab, a gene for resistance to the antibiotic ampicillin into a bacterial strain that is killed by ampicillin was introduced. If the susceptible bacteria incorporate the foreign DNA, they will become ampicillin resistant. The bacterium we use in our laboratory activity is Escherichia coli, which has been grown in a Petri dish on Luria Broth (LB) agar. Each colony in the Petri dish is made up of millions of individual cells. Escherichia coli is the most common bacterium in the human gut. It has been extensively studied in the laboratory and is an important research organism for molecular biology. E. coli reproduce very rapidly; a single microscopic cell can divide to form a visible colony with millions of cells overnight. Like all bacteria, E. coli has no nuclear envelope surrounding the bacterial chromosome and thus no true nucleus. All of the genes required for basic survival and reproduction are found in the ingle
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6a lab - Dhaval Rana Ap Biology December 16, 2009 Lab 6a:...

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