Lambda phage killing: infection and “parts” N Kuldell for Spring 2009

Heidelburg iGEM 2008: ecolicense to kill Part 2: DNA transfer, genetically programmed self-assembly and “parts” Background information

DNA transfer natural context: 3 mechanisms

“Transduction” by bacteriophage Protein coat encapsulates nucleic acids Bacteriophage: viruses that infect bacteria INFECT REPLICATE ASSEMBLE RELEASE

“Temperate” bacteriophage lambda INFECT REPLICATE ASSEMBLE RELEASE lytic

INFECT REPLICATE ASSEMBLE RELEASE lytic lysogenic “Temperate” bacteriophage lambda

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?

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?

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?

Genetically programmed infection iGEM context: phage infection via conjugation

Genetically programmed infection iGEM context: phage infection via conjugation 572/L18/L18.htm io.uwinnipeg. ca

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

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

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?

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?

Genetically programmed bi-stable switch natural context: epigenetic regulation Genes Dev. (2004)18(17): 2086–2094. doi: /gad lysogeny lysis

Genetically programmed bi-stable switch iGEM context: flip and hold in one state Genes Dev. (2004)18(17): 2086–2094. doi: /gad lysogeny promoterRBScI ORF transcriptional terminator

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?

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?

Fluorescence vs bioluminescence AI-2 AI-1

Fluorescence vs bioluminescence AI-2 AI-1

Fluorescence vs bioluminescence AI-2 AI-1

One last thing about fluorescence

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

Predator cell stable lysogen that infects by conjugation + CamR + GFP to follow process