Lambda phage killing:
infection and “parts”
N Kuldell for 20.020
Spring 2009
Heidelburg iGEM 2008:
ecolicense to kill
Part 2: DNA transfer, genetically
programmed self-assembly and “parts”
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Background information
DNA transfer
natural context: 3 mechanisms
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http://www.scq.ubc.ca/wp-content/uploads/2006/08/HorizontalTransfer.gif
“Transduction” by bacteriophage
Bacteriophage: viruses
that infect bacteria
Protein coat
encapsulates
nucleic acids
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INFECT
REPLICATE
RELEASE
ASSEMBLE
www.phagebiotech.com/
“Temperate” bacteriophage lambda
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INFECT
lytic
REPLICATE
RELEASE
ASSEMBLE
http://escience.ws/b572/L17/L17.htm
“Temperate” bacteriophage lambda
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INFECT
lytic
REPLICATE
RELEASE
lysogenic
ASSEMBLE
http://escience.ws/b572/L17/L17.htm
“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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INFECT
REPLICATE
RELEASE
ASSEMBLE
http://escience.ws/b572/L17/L17.htm
“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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INFECT
REPLICATE
RELEASE
ASSEMBLE
http://escience.ws/b572/L17/L17.htm
“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?
QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
INFECT
REPLICATE
RELEASE
ASSEMBLE
http://escience.ws/b572/L17/L17.htm
Genetically programmed infection
iGEM context: phage infection via conjugation
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Genetically programmed infection
iGEM context: phage infection via conjugation
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io.uwinnipeg.
ca
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http://escience.ws/b
572/L18/L18.htm
Genetically programmed infection
iGEM context: phage infection via conjugation
What “parts” are needed
Pilus….
“F’”
Origin for transfer…
“oriT”
Selectable marker… “CamR”
“Part” is a genetically-encoded,
human defined function
Model predicts: 10 killer cells kill
10^9 prey cells “in silico”
CamR
oriT
F’
Genetically programmed infection
iGEM context: phage infection via conjugation
What “parts” are needed
Pilus….
“F’”
Origin for transfer…
“oriT”
CamR
I714030
Selectable marker… “CamR”
“Part” is a genetically-encoded,
human defined function
oriT
F’
“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?
QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
INFECT
REPLICATE
RELEASE
ASSEMBLE
http://escience.ws/b572/L17/L17.htm
“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?
QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
INFECT
REPLICATE
RELEASE
ASSEMBLE
http://escience.ws/b572/L17/L17.htm
Genetically programmed bi-stable switch
natural context: epigenetic regulation
lysogeny
lysis
Genes Dev. (2004)18(17): 2086–2094.
doi: 10.1101/gad.1226004.
Genetically programmed bi-stable switch
iGEM context: flip and hold in one state
promoter
RBS
cI ORF
transcriptional
terminator
lysogeny
Genes Dev. (2004)18(17): 2086–2094.
doi: 10.1101/gad.1226004.
“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?
QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
INFECT
REPLICATE
RELEASE
ASSEMBLE
http://escience.ws/b572/L17/L17.htm
“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?
QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
INFECT
REPLICATE
RELEASE
ASSEMBLE
http://escience.ws/b572/L17/L17.htm
Fluorescence
vs bioluminescence
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AI-1
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Fluorescence
vs bioluminescence
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http://www.lifesci.ucsb.edu/~biolum/chem/
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Fluorescence
vs bioluminescence
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http://www.lifesci.ucsb.edu/~biolum/chem/
AI-1
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AI-2
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”
CamR
oriT
F’
Predator cell
stable lysogen that infects by conjugation
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+ CamR
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