Bacterial Relay Race 09 An illustration of a cell-to-cell - - PowerPoint PPT Presentation

Bacterial Relay Race ‘09

An illustration of a cell-to-cell communication device The TUDelft iGEM team presents:

Calin, Daniel, Saeed, Sriram, Tim and Tim

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The TUDelft iGEM team

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Cell-to-cell communication

• Principles of cellular communication
• Propagation of a set of instructions
• Bacterial relay race
• Conjugation system
• Self-destructive plasmid
• Time-delay device
• Quorum sensing is limited to binary signals

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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System Overview

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Conjugation

• Stop entry exclusion
• Knockout trbK
• Stop conjugative transfer of

helper plasmid

• Knockout oriTR
• Address lethal zygosis
• Knockout trbC
• λ-red knockouts

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Conjugation

Conjugation testing plasmids

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Conjugation

Entry exclusion expression blocks incoming transfers

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Modeling

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Modeling

Two modes of propagation

• 1. Conjugation
• 8 to 20 μm / hr
• 2. Cell division
• > 20 μm / hr

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Self-destructive plasmid

• IPTG inducible destruction
• I-SceI homing endonuclease
• 30 bp cut site
• Induction stops GFP expression

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Self-destructive plasmid

• Cut site biobricked
• Successful assembly
• In vivo testing unsuccessful
• Undocumented

degradation tag

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Time-delay device

• Two approaches:
• synthetic transcriptional cascade
• biosynthetic AND gate

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Time-delay device

Two level negative cascade

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Modeling the Time-delay

• Stability
• Sensitivity
• Design

Recommendations

too short infinite delay AWESOME!

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Time-delay device

• Plasmids assembled successfully
• Tunable delay for future teams
• Delay time agrees with predicted value

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Time-delay device

Key Lock

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Issues in synthetic biology

~ Building ~ Reductionism

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Concepts of life

Time

LIFE

traditional

LIFE??

scientific

“Research” Reductionism Bottom up Understanding? Building?

Questions Concepts Process

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Methods

• Literature review
• Survey
• 242 participants from

iGEM community

• Analysis
• Quantitative
• Qualitative
• Raise awareness on ethical issues
• Gain insight in opinions and perceptions

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Survey Results

Will the reductionist approach lead to an understanding of life?

No, living systems are too complex. No, biological phenomena are fundamentally special. Yes, we will eventually fully understand life.

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Survey results

For all results see: http://2009.igem.org/Team:TUDelft/Ethics

Agree

A complex living system is more than the sum of its biological components

Not Agree

Life is physical and can be explained materialistically

Agree Not Agree 65% 23% 13% 19% 40% 40%

Overview Conjugation Self-destruction Time-delay Ethics Achievements

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Major Achievements

• Engineered a tunable delay device
• Characterized and submitted 21 new BioBricks
• Characterized several exisiting BioBricks
• New toolbox for modeling conjugation systems
• Lock and Key generation algorithm
• An extensive study on Human Practice

Overview Conjugation Self-degradation Time-delay Ethics Achievements

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Acknowledgements

• Supervisors
• Emrah Nikerel
• Domenico Bellomo
• Janine Kiers
• Advisors
• Esengül Yıldırım
• Steven Flipse

Sponsors:

Bacterial Relay Race ‘09

An illustration of a cell-to-cell communication device For more information: http://2009.igem.org/Team:TUDelft

Calin, Daniel, Saeed, Sriram, Tim and Tim