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Lambda phage killing: infection and “parts” N Kuldell for 20.020 Spring 2009
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Heidelburg iGEM 2008: ecolicense to kill Part 2: DNA transfer, genetically programmed self-assembly and “parts” Background information
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DNA transfer natural context: 3 mechanisms http://www.scq.ubc.ca/wp-content/uploads/2006/08/HorizontalTransfer.gif
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“Transduction” by bacteriophage www.phagebiotech.com/ Protein coat encapsulates nucleic acids Bacteriophage: viruses that infect bacteria INFECT REPLICATE ASSEMBLE RELEASE
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“Temperate” bacteriophage lambda http://escience.ws/b572/L17/L17.htm INFECT REPLICATE ASSEMBLE RELEASE lytic
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http://escience.ws/b572/L17/L17.htm INFECT REPLICATE ASSEMBLE RELEASE lytic lysogenic “Temperate” bacteriophage lambda
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http://escience.ws/b572/L17/L17.htm INFECT REPLICATE ASSEMBLE RELEASE “Temperate” bacteriophage lambda Some great ??s What guides lytic/lysogenic decision? What keeps lysogen stable? What triggers lysogen to lytic cycle?
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http://escience.ws/b572/L17/L17.htm INFECT REPLICATE ASSEMBLE RELEASE “Temperate” bacteriophage lambda Some great ??s the Heidelberg team asked: Can we infect prey from lysogen (=predator)? Can we keep lysogen from lysing itself? Can we monitor lysis and lysogeny?
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http://escience.ws/b572/L17/L17.htm INFECT REPLICATE ASSEMBLE RELEASE “Temperate” bacteriophage lambda Some great ??s the Heidelberg team asked: Can we infect prey from lysogen (=predator)? Can we keep lysogen from lysing itself? Can we monitor lysis and lysogeny?
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Genetically programmed infection iGEM context: phage infection via conjugation
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Genetically programmed infection iGEM context: phage infection via conjugation http://escience.ws/b 572/L18/L18.htm io.uwinnipeg. ca
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Genetically programmed infection iGEM context: phage infection via conjugation What “parts” are needed Pilus…. “F’” Origin for transfer… “oriT” Selectable marker… “CamR” Model predicts: 10 killer cells kill 10^9 prey cells “in silico” F’ oriT CamR “Part” is a genetically-encoded, human defined function
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Genetically programmed infection iGEM context: phage infection via conjugation What “parts” are needed Pilus…. “F’” Origin for transfer… “oriT” Selectable marker… “CamR” F’ oriT CamR “Part” is a genetically-encoded, human defined function I714030
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http://escience.ws/b572/L17/L17.htm INFECT REPLICATE ASSEMBLE RELEASE “Temperate” bacteriophage lambda Some great ??s the Heidelberg team asked: Can we infect prey from lysogen (=predator)? Can we keep lysogen from lysing itself? Can we monitor lysis and lysogeny?
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http://escience.ws/b572/L17/L17.htm INFECT REPLICATE ASSEMBLE RELEASE “Temperate” bacteriophage lambda Some great ??s the Heidelberg team asked: Can we infect prey from lysogen (=predator)? Can we keep lysogen from lysing itself? Can we monitor lysis and lysogeny?
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Genetically programmed bi-stable switch natural context: epigenetic regulation Genes Dev. (2004)18(17): 2086–2094. doi: 10.1101/gad.1226004. lysogeny lysis
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Genetically programmed bi-stable switch iGEM context: flip and hold in one state Genes Dev. (2004)18(17): 2086–2094. doi: 10.1101/gad.1226004. lysogeny promoterRBScI ORF transcriptional terminator
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http://escience.ws/b572/L17/L17.htm INFECT REPLICATE ASSEMBLE RELEASE “Temperate” bacteriophage lambda Some great ??s the Heidelberg team asked: Can we infect prey from lysogen (=predator)? Can we keep lysogen from lysing itself? Can we monitor lysis and lysogeny?
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http://escience.ws/b572/L17/L17.htm INFECT REPLICATE ASSEMBLE RELEASE “Temperate” bacteriophage lambda Some great ??s the Heidelberg team asked: Can we infect prey from lysogen (=predator)? Can we keep lysogen from lysing itself? Can we monitor lysis and lysogeny?
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Fluorescence vs bioluminescence AI-2 AI-1
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Fluorescence vs bioluminescence AI-2 AI-1 http://www.lifesci.ucsb.edu/~biolum/chem/
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Fluorescence vs bioluminescence AI-2 AI-1 http://www.lifesci.ucsb.edu/~biolum/chem/
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One last thing about fluorescence
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Genetically programmed infection iGEM context: phage infection via conjugation What “parts” are needed Pilus…. “F’” Origin for transfer… “oriT” Selectable marker… “CamR” Stable lysogen… “cI” Moniter… “GFP” F’ oriT CamR
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Predator cell stable lysogen that infects by conjugation + CamR + GFP to follow process
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