2007 brown igem team n.
Download
Skip this Video
Loading SlideShow in 5 Seconds..
2007 Brown iGEM Team PowerPoint Presentation
Download Presentation
2007 Brown iGEM Team

Loading in 2 Seconds...

play fullscreen
1 / 46

2007 Brown iGEM Team - PowerPoint PPT Presentation


  • 120 Views
  • Uploaded on

2007 Brown iGEM Team. 7 undergraduates 7 grad student advisors 2 Faculty advisors 9 faculty sponsors. 1/45. Brown iGEM. international genetically engineered machines competition. June Update. 2/45. What is iGEM?. Biology Engineering Standardization. 3/45. Science - Nuts and Bolts.

loader
I am the owner, or an agent authorized to act on behalf of the owner, of the copyrighted work described.
capcha
Download Presentation

PowerPoint Slideshow about '2007 Brown iGEM Team' - candy


An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.


- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript
2007 brown igem team
2007 Brown iGEM Team
  • 7 undergraduates
  • 7 grad student advisors
  • 2 Faculty advisors
  • 9 faculty sponsors

1/45

brown igem
Brown iGEM

international genetically engineered machines competition

June Update

2/45

what is igem
What is iGEM?
  • Biology
  • Engineering
  • Standardization

3/45

science nuts and bolts
Science - Nuts and Bolts
  • Standardizing biology
  • Systematic engineering
  • Apply biological technology

4/45

previous projects
Previous Projects
  • Bacterial Photo Film - U. Texas (published in Nature)

5/45

previous projects1
Previous Projects
  • Sepsis Treatment - Slovenia
  • Banana/Wintergreen Smelling Cells - MIT
  • Arsenic Water Detection - Edinburgh

5/45

slide7
Anderson, J. C., Clarke, E. J., Arkin, A. P., & Voigt, C. A. (2005) Environmentally Controlled Invasion of Cancer Cells by Engineered Bacteria, Journal of Molecular Biology

6/45

slide8
Anderson, J. C., Clarke, E. J., Arkin, A. P., & Voigt, C. A. (2005) Environmentally Controlled Invasion of Cancer Cells by Engineered Bacteria, Journal of Molecular Biology

Aerobic Conditions

Low Cell Density

>0.02% Arabinose

OFF

Hypoxia

High Cell Density

<0.02% Arabinose

ON

Inv

induction

INVASION

7/45

undergraduate teams
Undergraduate Teams

+ Imagined

+ Designed

+ Implemented

by a team of undergraduates

8/45

progress
Progress
  • Brown’s 2nd year
  • UTRA grants
  • Lab space in Multi Disciplinary Lab
  • Equipment sponsorship

9/45

why do we need this
Why do we need this?
  • Public health concern: Lead in soil, paint, water, dust

11/45

why do we need this1
Why do we need this?
  • Public health concern: Lead in soil, paint, water, dust
  • Lead Poisoning is often caused by ingesting contaminated drinking water, or soil. It can cause neurological and gastrointestinal disorders, especially among children.

12/45

why do we need this2
Why do we need this?
  • Public health concern: Lead in soil, paint, water, dust
  • Lead Poisoning is often caused by ingesting contaminated drinking water, or soil. It can cause neurological and gastrointestinal disorders, especially among children.
  • The legal limit of lead in drinking water is 15 parts per billion.

13/45

why do we need this3
Why do we need this?

Current ways of testing for lead either require expensive chemical lab analysis or involve inaccurate home kits.

So why do we want a biosensor?

- Cheap

- Sensitive

- Quick

- Specific

14/45

slide16

We have spliced together biobrick parts into plasmids in E. Coli. The bacteria express our genetic devices to create a lead detector.

15/45

slide17

General Design

Lead

Lead Detection

Lead Detection

Signal Amplification

Signal Amplification

Fluorescent Output

16/45

2 system components
2 System Components

Part 1: Lead Detection

Part 2: Signal Amplification

18/45

slide20

Part 1: Lead Detection

Lead Receptor Protein

Lead Receptor Gene

Always On

19/45

slide21

Part 1: Lead Detection

Lead

Activated Lead Receptor Protein

Lead Receptor Protein

Lead Receptor Gene

Always On

20/45

slide22

Part 1: Lead Detection

Lead

Activated Lead Receptor Protein

Lead Receptor Protein

Activated Lead Receptor Protein

Lead Receptor Gene

Always On

Signal Amplification promoter

21/45

slide23

Part 1: Lead Detection

  • This is sensitive ONLY to lead; other metals will not activate it.

22/45

Chen P, Greenberg B, Taghavi S, Romano C, van der Lelie D, He C (2005) An exceptionally selective lead(II)-regulatory protein from Ralstonia metallidurans: development of a fluorescent lead(II) probe. Angew Chem Int Ed Engl 44:2715–2719

slide24

General Design

Lead

Lead Detection

Lead Detection

Signal Amplification

Signal Amplification

Fluorescent Output

23/45

slide25

Part 2: Signal Amplification

Activated Lead Receptor Protein

Signal Amplification promoter

24/45

slide26

Part 2: Signal Amplification

Message

Activated Lead Receptor Protein

Message Producer Gene

Signal Amplification promoter

25/45

slide27

Part 2: Signal Amplification

Message Producer Gene

Signal (GFP)

Message

Activated Lead Receptor Protein

Message Producer Gene

Signal Amplification promoter

26/45

slide28

Part 2: Signal Amplification

Message Producer Gene

Signal (GFP)

Message

Activated Lead Receptor Protein

Repeated activation of this promoter causes amplification of the signal.

Message Producer Gene

Signal Amplification promoter

27/45

3 system components
3 System Components

Lead Detection

1. Detector Sequence

2. Leakiness Filter to Eliminate False Positives

Signal Amplification

3. Positive Feedback Loop for Amplification

We’ve added a “Leakiness Filter” as an additional component to the system. This gives our circuit one more level of complexity. However, the black box diagram is generally the same.

28/45

slide30

NO LEAD

Lead Activator

Message Activator

Always On

Transcription factors are constitutively made by the first promoter.

Message Maker

Filter Repressor

Lead Detector

Message Destroyer

Filter

Filter Eliminates Stray Message Molecules to Prevent False Feedback Loop Activation

These proteins are poised to activate the Lead Detector promoter and Message Receiver promoter upon addition of lead.

Stray Message Molecules Can Falsely Activate the Feedback Loop.

Message Maker

GFP Reporter

Message Receiver

29/45

slide31

Lead Activator

Feedback Activator

Always On

Message Maker

Filter Repressor

Lead turns on Detector promoter

Lead Detector

Message Destroyer

Leakiness Filter promoter gets turned off

Filter

+

Fluorescent Protein Output

Message Maker

GFP Reporter

Message Receiver

30/45

slide32

Lead Activator

Feedback Activator

Always On

Message Maker

Filter Repressor

1

Lead Detector

Message Destroyer

2

Filter

Message Maker

GFP Reporter

Message Receiver

3

31/45

slide33

PbrR

LuxR

pTet

LuxI

LacI

1

Pbr

aiiA

2

pLac

LuxI

GFP

pLux

32/45

3

how this project advances science
How this project advances science:
  • Characterization of Existing Parts
  • Adding New Parts and Devices
    • Lead Promoter and Transcription Factor
    • Amplifier Device
  • Generalize to future biosensors
    • Arsenic, Cadmium, Mercury, Zinc

34/45

what is tri stability
What is Tri-stability?
  • A tri-stable switch has three distinct and inducible states

A

B

C

36/45

achieving tri stability
Achieving Tri-stability

Input A

Output A

State A

Input B

Output B

State B

Input C

Output C

State C

37/45

slide39

The Architecture

Arabinose

RFP

pBad/Ara

tetR

lacI

IPTG

CFP

araC

tetR

pLac

Tetracycline

YFP

pTet

lacI

araC

38/45

characterization
Characterization
  • Characterization is an essential aspect of iGEM
  • It is a step towards standardization - giving others all the details needed to use the part.

39/45

slide41

Uses

  • Differentiation of stem cells
  • Turn on/off three different proteins in cell
  • Cellular logic
  • Tissue Engineering

40/45

timeline
Timeline
  • Now:
    • Start cloning
    • PCR lead promoter
    • Clone tri-stable switch
    • Characterize parts
    • Test systems
    • Send back to the Registry
  • August: End of lab work
  • November: Jamboree at MIT

41/45

why brown
Why Brown?
  • Innovators
  • Entrepreneurs
  • A great place for new ideas!

42/45

the future
The Future
  • Educate others about iGEM and synthetic biology
  • Synthetic Biology Course Offering in Fall 07! Led by Prof. Gary Wessel

43/45

special thanks to
Special Thanks To:

Office of the Dean of the CollegeOffice of the PresidentThe Atlantic PhilanthropiesThe Center for Computational and Molecular BiologyDepartment of PhysicsEngineering DepartmentDepartment of Molecular Biology, Cell Biology, and BiochemistryDepartment of Molecular Pharmacology, Physiology, and BiotechnologyThe Multi Disciplinary LabPfizerLabnetNanodrop

44/45