First, about our team: Ali Celentano Dominique MacCalla Nicole - - PowerPoint PPT Presentation

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First, about our team: Ali Celentano Dominique MacCalla Nicole - - PowerPoint PPT Presentation

May 20, 2023 235 likes •1.42k views

First, about our team: Ali Celentano Dominique MacCalla Nicole Matamala Danielle Peters Courtney Pozzi Niteesh Sundaram Lena Wang Ali Celentano: Senior, Business/Biology Dominique MacCalla: Nicole Matamala Danielle Peters Courtney Pozzi

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First, about our team:

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Ali Celentano Dominique MacCalla Nicole Matamala Danielle Peters Courtney Pozzi Niteesh Sundaram Lena Wang

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Ali Celentano: Senior, Business/Biology Dominique MacCalla: Nicole Matamala Danielle Peters Courtney Pozzi Niteesh Sundaram Lena Wang

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Ali Celentano: Senior, Business/Biology Dominique MacCalla: Junior, MatSci/BME Nicole Matamala Danielle Peters Courtney Pozzi Niteesh Sundaram Lena Wang

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Ali Celentano: Senior, Business/Biology Dominique MacCalla: Junior, MatSci/BME Nicole Matamala: Sophomore, CivE/BME Danielle Peters Courtney Pozzi Niteesh Sundaram Lena Wang

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Ali Celentano: Senior, Business/Biology Dominique MacCalla: Junior, MatSci/BME Nicole Matamala: Sophomore, CivE/BME Danielle Peters: Senior, Biology Courtney Pozzi Niteesh Sundaram Lena Wang

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Ali Celentano: Senior, Business/Biology Dominique MacCalla: Junior, MatSci/BME Nicole Matamala: Sophomore, CivE/BME Danielle Peters: Senior, Biology Courtney Pozzi: Soph, Industrial

Design/BME

Niteesh Sundaram Lena Wang

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Ali Celentano: Senior, Business/Biology Dominique MacCalla: Junior, MatSci/BME Nicole Matamala: Sophomore, CivE/BME Danielle Peters: Senior, Biology Courtney Pozzi: Soph, Industrial Design,

BME

Niteesh Sundaram: Senior, ECE Lena Wang

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Ali Celentano: Senior, Business/Biology Dominique MacCalla: Junior, MatSci/BME Nicole Matamala: Sophomore, CivE/BME Danielle Peters: Senior, Biology Courtney Pozzi: Soph, Industrial

Design/BME

Niteesh Sundaram: Senior, ECE Lena Wang: Senior, Biology/BME

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Ali Celentano: Senior, Business/Biology Dominique MacCalla: Junior, MatSci/BME Nicole Matamala: Sophomore, CivE/BME Danielle Peters: Senior, Biology Courtney Pozzi: Soph, Industrial

Design/BME

Niteesh Sundaram: Senior, ECE Lena Wang: Senior, Biology/BME

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Our Project

The development of a biosensor for estrogenic activity in water

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The Problem: Endocrine disruptors, such

as estrogens find their way into our water systems and are incredibly hard to detect.

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Between 1999 and 2000, the U.S. Geological Survey sampled 139 surface waters throughout the U.S. Through this study, it was determined that 80% of those waters contained endocrine disrupting chemicals, most of them being estrogens.

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Environmental Impact

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Environmental Impact

Fish Feminization

  • Studies have documented crashing fish

populations due to the feminization of male fish by estrogens.

  • Entire ecosystems are affected.
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How do we demonstrate ecosystem effects?

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Modeling allows us to simulate possible scenarios to predict the effect of estrogenic activity on ecosystems.

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Elements of Environmental NetLogo Model

Bird Population Fish Population

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No Estrogen

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Low Estrogen

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Medium Estrogen

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High Estrogen

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How does modeling inform policy?

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The Politics of Regulation

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The Politics of Regulation

Thousands of estrogen-containing products enter our water system and are difficult (pharmaceuticals) or impossible (natural estrogens) to regulate.

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Current Policies

  • Switzerland is investing in source

control regulations for estrogenic products.

  • EU wants less expensive

technologies that detect endocrine disruptors.

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Current Policies

  • Strategy Micropoll Project has shown

that pollutants in surface waters can be reduced by at least 80% with additional treatment of already cleaned wastewater.

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Current Policies

  • Amendment to the Swiss Water

Protection Law: upgrading of 100

  • f ~700 Swiss wastewater treatment

plants, requiring an investment of approximately 1.2 billion Swiss francs (1.247 billion USD).

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Current Policies

  • "Polluter pays” wastewater tax:

maximum tax of 9 Swiss francs (9.35 USD) per person per year collected until

  • 2040. The tax should finance 75% of the

investment costs.

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Our Solution: T

  • engineer a rapid

sensitive biosensor to detect estrogenic activity in water.

Responding to Policy

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Industrial Impact: Our biosensor is

intended to improve on the current industrial standard, which takes days to complete.

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Synthesis of STREAM

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We began by analyzing a set of fluorescent proteins in order to determine the optimal reporter for our biosensor.

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Fluorescent Protein Evaluation

  • Analyzed five fluorescent proteins

as reporters for our sensor

  • Technology: TECAN plate reader
  • Measurement: signal-to-noise ratios
  • Cell Lines: MACH and TOP10 E. coli
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Fluorescent Protein Evaluation

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Fluorescent Protein Evaluation

Blue

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Fluorescent Protein Evaluation

Blue Green

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Fluorescent Protein Evaluation

Blue Green Yellow

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Fluorescent Protein Evaluation Orange

Blue Green Yellow

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Fluorescent Protein Evaluation Orange

Red Blue Green Yellow

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Fluorescent Protein Results

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Our Biosensor

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Bacterial Biosensor Model

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Characterizing Our Biosensor

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Modeling Verification

Using a rule-based model helped save time in the wet lab and identify bottlenecks

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Modeling Verification

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Modeling Verification

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BioBrick Improvements

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BioBrick Improvements

  • Codon optimization to improve yield
  • Monomeric version
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KillerRed and SuperNova

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KillerRed and SuperNova

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iGEM Measurement Interlab Study

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iGEM Measurement Interlab Study

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Collaborations

  • iGEM Measurement Interlab Study
  • IvyT

ech Kit Modification

  • iGEM Meetup
  • Pitt Modeling
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Collaborations

IvyT ech Wheat Germ DNA Extraction

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5 teams, 50 iGEMers

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 Addressed current policy dealing with endocrine disruptors  Used modeling simulations to demonstrate impact  Applied our backgrounds to teaching the community about synthetic biology  Ensured that our practices will last longer than one iGEM summer

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 Addressed current policy dealing with endocrine disruptors  Used modeling simulations to demonstrate impact  Applied our backgrounds to teaching the community about synthetic biology  Ensured that our practices will last longer than one iGEM summer

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Our project stemmed out of current policy needs in:

  • European Union
  • Switzerland
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Policy into Practice

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Who’s at home?

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Who’s at home? Underprivileged elementary and middle school students who just started learning about science in Pittsburgh, PA

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How does policy help them understand

  • ur project?
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How does policy help them understand

  • ur project?

It really doesn’t.

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What practices do we develop to engage, educate, and excite students about synthetic biology?

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What practices do we develop to engage, educate, and excite students about synthetic biology? Can these practices last longer than an IGEM summer?

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 Addressed current policy dealing with endocrine disruptors  Used modeling simulations to demonstrate impact  Applied our backgrounds to teaching the community about synthetic biology  Ensured that our practices will last longer than one iGEM summer

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Environmental NetLogo Simulation

helped us visually explain environmental impact of estrogens motivated us to work on this project

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 Addressed current policy dealing with endocrine disruptors  Used modeling simulations to demonstrate impact  Applied our backgrounds to teaching the community about synthetic biology  Ensured that our practices will last longer than one iGEM summer

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Friday June 6, 2014 1 PM “This sounds like a great opportunity! It would be best for us if you could do three 80 minute classes for scheduling purposes.” –Hilary Buttenfield, ECS

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Engineering + Design In a Synthetic Biology World?

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SynBio Educational Series

The iGEM team created and aided in the development of four kits for students and teachers to use in a classroom setting

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SynBio Educational Series

The iGEM team created and aided in the development of four kits for students and teachers to use in a classroom setting Creature Feature DNA Base Pairing Kit Wheat Germ DNA Extraction Strawberry DNA Extraction

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iGEM asked us to beyond the bench, We decided to go beyond the bench[mark]

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Creature Feature An engaging modeling lab that takes foundational concepts from genetics and evolution and concludes into the field of synthetic biology.

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DNA Base Pairing Kit: (improved) DNA modeling kit for the visualizing the principles of base pairing.

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Wheat Germ and Strawberry DNA Extraction

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Wheat Germ and Strawberry DNA Extraction

Improvements

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Wheat Germ and Strawberry DNA Extraction

Improvements Wheat Germ Strawberry

Self contained activity Reduced time to 8 minutes Simplified Able to store activity infinitely Easier Introduced a syringe filtering technique Reduced time to10 minutes Created much higher DNA yields Less mess Easier

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Future Initiative

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Future Initiative

  • Beta with even more students
  • Taught teachers who
  • Gave feedback
  • Provided support
  • Demanded lesson plans immediately
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Future Initiative

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Future Initiative

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Outreach by Numbers

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Outreach by Numbers

Reached 655 people; 615 students 40 educators 648 labs completed to date

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Outreach by Numbers

Reached 655 people; 615 students 40 educators 648 labs completed to date After the jamboree?

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1080+

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Outreach by Location

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Project Conclusions

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Biological

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Biological

  • Our E. coli biosensor platform showed a 400

fold increase in RFP signal but was unresponsive to estrogen.

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Biological

  • Our E. coli biosensor platform showed a 400

fold increase in RFP signal but was unresponsive to estrogen.

  • The Mycbacterium tuberculosis RecA intein

will be tested (Skretas and Wood 2005) and

  • ther inteins that splice optimally at 37oC

(Shah and Muir, 2014)

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Outreach

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Outreach

  • All of our kits and labs will be available online
  • n DNAZone starting January 2015.
  • Creature Feature has 20 more scheduled and

confirmed appearances by December for approximately1080 people

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Acknowledgements

  • Dr. Cheryl Telmer, main advisor
  • Dr. Natasa Miskov-Zivanov, modeling advisor
  • Dr. Marcel Bruchez, advisor, providing funding
  • Dr. Diana Marculescu, advisor, securing funding
  • Dr. Carrie Doonan, outreach advisor
  • Dr. Jeanne VanBriesen, environmental advice
  • Dr. Terrence Collins, advice on policies
  • Dr.

Catalina Achim and Hannah Diorio-Toth, DNAzone

  • Taylor Canady and Genoa Warner, DNA kits
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Acknowledgements cont.

  • Annette Jacobson and Nathan Urban, funding
  • Jason Lohmueller, experimental advice
  • James Faeder, BioNetGen advice
  • Yi Wang, provided competent cells
  • Jonathan Dewerd, wiki advice
  • Eric Pederson, past iGEM team member, advice
  • Yang Choo, past iGEM team member, advice
  • David Casillas and Carol Lee, Biology Business
  • ffice
  • Amie Rose DiTomasso, Sciencepalooza coordinator
  • Matthew Salyers, Web designer, Biological Sciences
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Sponsors

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