Population_Control_Gene_Circuit_Week_Two - POPULATION...

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POPULATION CONTROL GENE CIRCUIT: WEEK TWO Safety: The Escherichia coli bacteria used in this lab have been modified to be non-virulent, but may induce an acute immune response in a very small percentage of individuals (this may occur whenever one is exposed to foreign materials or organisms). You must take the following safety precautions: Wash your hands when entering and before exiting the lab Wear a lab coat and gloves (dispose of after the lab) No eating or drinking in the lab Wear closed toe shoes Objectives: In this lab, you will: Week 1 – transform “population control” gene circuit plasmid into E. coli Week 2 - analyze growth of bacteria containing the “population control” gene circuit Background: Population Control Gene Circuit (Lab Week 2) When normal bacteria are introduced in a fixed resource environment in the wild or in the laboratory (i.e in growth media with suitable conditions such as temperature, pH, oxygen, and nutrients), cells grow in four phases: lag, log, stationary, and death. 1. Lag phase. Cells grow relatively slowly as they acclimate to the growth conditions and available resources. 2. Log (logarithmic) phase. While resources are plentiful, population density rises exponentially with time. The cycle continues so that a single parent cell is the progenitor of an exponentially increasing number of daughter cells: 2, 4, 8, 16, 32, 64, etc. In E. coli, each cell cycle takes approximately 20-60 minutes. 3. Stationary phase. As resources are depleted, cells compete for limited resources. The total number of cells becomes relatively stable, as the number of cells that are formed is approximately equal to the number of cells that die. 4. Death phase. Once resources have been depleted and toxic waste products have built up, the environment is inhospitable for cell life. (image modified from http://www.scienceaid.co.uk/biology/microorganisms/populations.html) Time Cell Density
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With the naked eye, cell growth can be estimated by “turbidity,” or the increasingly cloudy appearance of growth media as the cell population increases. An accurate and simple method of measuring cell growth is by observing the absorbance of light (600 nm λ ). In this lab, cell growth will be determined by absorbance measurements taken at regular intervals (30 minutes) by an automated instrument, the Victor3 plate reader. Rather than the simple growth curve described above, the growth of the E. coli population you will study in this lab will oscillate as it “monitors” its own population density. This is done through a process called quorum sensing, which enables bacteria to coordinate their behavior by communicating with each other through signaling
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This note was uploaded on 08/03/2009 for the course BME 100 taught by Professor Yuan during the Spring '07 term at Duke.

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Population_Control_Gene_Circuit_Week_Two - POPULATION...

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