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Lambda phage killing: infection and “parts” N Kuldell for 20.020 Spring 2009

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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”. Background information. DNA transfer natural context: 3 mechanisms. - PowerPoint PPT Presentation

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Page 1: Lambda phage killing: infection and “parts”

Lambda phage killing: infection and “parts”

N Kuldell for 20.020

Spring 2009

Page 2: Lambda phage killing: infection and “parts”

Heidelburg iGEM 2008: ecolicense to kill

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Part 2: DNA transfer, genetically programmed self-assembly and “parts”

Background information

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Page 3: Lambda phage killing: infection and “parts”

DNA transfer

natural context: 3 mechanisms

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http://www.scq.ubc.ca/wp-content/uploads/2006/08/HorizontalTransfer.gif

Page 4: Lambda phage killing: infection and “parts”

“Transduction” by bacteriophage

www.phagebiotech.com/

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Protein coat encapsulates nucleic acids

Bacteriophage: viruses that infect bacteria

INFECT

REPLICATE

ASSEMBLE

RELEASE

Page 5: Lambda phage killing: infection and “parts”

“Temperate” bacteriophage lambda

http://escience.ws/b572/L17/L17.htm

INFECT

REPLICATE

ASSEMBLE

RELEASE

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lytic

Page 6: Lambda phage killing: infection and “parts”

http://escience.ws/b572/L17/L17.htm

INFECT

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lytic

lysogenic

“Temperate” bacteriophage lambda

Page 7: Lambda phage killing: infection and “parts”

http://escience.ws/b572/L17/L17.htm

INFECT

REPLICATE

ASSEMBLE

RELEASE

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“Temperate” bacteriophage lambda

Some great ??s

What guides lytic/lysogenic decision?

What keeps lysogen stable?

What triggers lysogen to lytic cycle?

Page 8: Lambda phage killing: infection and “parts”

http://escience.ws/b572/L17/L17.htm

INFECT

REPLICATE

ASSEMBLE

RELEASE

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“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?

Page 9: Lambda phage killing: infection and “parts”

http://escience.ws/b572/L17/L17.htm

INFECT

REPLICATE

ASSEMBLE

RELEASE

QuickTime™ and aTIFF (Uncompressed) decompressor

are needed to see this picture.

“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?

Page 10: Lambda phage killing: infection and “parts”

Genetically programmed infection iGEM context: phage infection via conjugation

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Page 11: Lambda phage killing: infection and “parts”

Genetically programmed infection iGEM context: phage infection via conjugation

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http://escience.ws/b572/L18/L18.htm

io.uwinnipeg.ca

Page 12: Lambda phage killing: infection and “parts”

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

Page 13: Lambda phage killing: infection and “parts”

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

http://partsregistry.org/wiki/index.php/Part:BBa_I714030
Page 14: Lambda phage killing: infection and “parts”

http://escience.ws/b572/L17/L17.htm

INFECT

REPLICATE

ASSEMBLE

RELEASE

QuickTime™ and aTIFF (Uncompressed) decompressor

are needed to see this picture.

“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?

Page 15: Lambda phage killing: infection and “parts”

http://escience.ws/b572/L17/L17.htm

INFECT

REPLICATE

ASSEMBLE

RELEASE

QuickTime™ and aTIFF (Uncompressed) decompressor

are needed to see this picture.

“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?

Page 16: Lambda phage killing: infection and “parts”

Genetically programmed bi-stable switch natural context: epigenetic regulation

Genes Dev. (2004)18(17): 2086–2094. doi: 10.1101/gad.1226004.

lysogeny

lysis

Page 17: Lambda phage killing: infection and “parts”

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

promoter RBS cI ORF transcriptional terminator

Page 18: Lambda phage killing: infection and “parts”

http://escience.ws/b572/L17/L17.htm

INFECT

REPLICATE

ASSEMBLE

RELEASE

QuickTime™ and aTIFF (Uncompressed) decompressor

are needed to see this picture.

“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?

Page 19: Lambda phage killing: infection and “parts”

http://escience.ws/b572/L17/L17.htm

INFECT

REPLICATE

ASSEMBLE

RELEASE

QuickTime™ and aTIFF (Uncompressed) decompressor

are needed to see this picture.

“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?

Page 20: Lambda phage killing: infection and “parts”

QuickTime™ and aTIFF (Uncompressed) decompressor

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Fluorescence vs bioluminescence

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AI-2AI-1

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Page 21: Lambda phage killing: infection and “parts”

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Fluorescence vs bioluminescence

AI-2AI-1

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http://www.lifesci.ucsb.edu/~biolum/chem/

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Page 22: Lambda phage killing: infection and “parts”

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Fluorescence vs bioluminescence

AI-2AI-1

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http://www.lifesci.ucsb.edu/~biolum/chem/

Page 23: Lambda phage killing: infection and “parts”

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One last thing about fluorescence

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Page 24: Lambda phage killing: infection and “parts”

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

Page 25: Lambda phage killing: infection and “parts”

Predator cellstable lysogen that infects by conjugation

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+ CamR

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