Synthetic Biology for Everyday Life Our Project F ats ils rease - - PowerPoint PPT Presentation

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Synthetic Biology for Everyday Life Our Project F ats ils rease - - PowerPoint PPT Presentation

Apr 07, 2023 273 likes •603 views

Synthetic Biology for Everyday Life Our Project F ats ils rease s O G INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT Hayton Wastewater Treatment Facility 14 tonne fatberg found in Kingston,

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“Synthetic Biology for Everyday Life”

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

14 tonne fatberg found in Kingston, London, UK

Our Project

Hayton Wastewater Treatment Facility

ats F ils O rease G

150,000 major blockages caused by FOGs every year in the UK.

s

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Hence…

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

SocioBricks

a background and basis for the project work: from social research to

  • utreach

to ensure projects are conducted ethically under legislation in different countries strategies to accomplish the work's goals: be they answering a social question to public engagement quantitative or qualitative, relating back to the initial framework and the methods employed to analyse the findings the communication of the work done can be conveyed in several

  • forms. In this collection, the user can find examples effective outputs

from previous iGEM teams

Framework

Ethics Methods Analysis Outputs

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

SocioBricks

Framework Ethics Methods Output

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

SocioBricks

Framework Methods Analysis Output Sheffield 2010 Sheffield 2014

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Problem: Fatbergs

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Initial Design

Keratin FAT

Fat Breakdown Hair Breakdown

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Initial Design

Criteria

  • Secrete enzyme to degrade fat  TliA

lipase via type I secretion

  • Secrete enzyme to degrade hair 

KerUS Keratinase (self-secretion)

  • Respond to fat levels  PfadR promoter
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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Initial Design

Criteria

  • Secrete lipid responsive lipase and

keratinase enzymes

LIPID RESPONSIVE KERATINASE

PfadR RBS kerUS Terminators BBa_K661060 (BBa_K1498000) PfadR RBS tliA Lipase Terminators BBa_K861060 BBa_K258006

PtetR RBS PrtD Domain Terminator RBS PrtE Domain Terminator RBS PrtF Domain Terminators BBa_R0040 B0034 K258100 B0010 B0034 K258101 B0010 B0034 K258102 B0010, B0012

LIPID RESPONSIVE LIPASE (BBa_K1498001) TYPE I SECRETION MODULE (BBa_K258008)

Criteria

  • Secrete lipid responsive lipase and

keratinase enzymes

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

FadR Characterisation Bba_K861061

TECAN data showing fluorescence produced from the fat responsive promoter / constitutive promoter over a 16 hour time period.

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

FadR Characterisation Bba_K861061

TECAN data showing fluorescence produced from the fat responsive promoter / constitutive promoter over a 16 hour time period.

  • 4 different concentration of oleic acid
  • No significant response to oleic acid by

the promoter

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

FadR Characterisation

Constitutive RFP fadR promoter- RFP fadR promoter

Promoter shows no response to fat levels

  • Cells DAPI blue stained

to show placement

  • Grown in 2% Oleic Acid

for 24 hours

  • RFP excited using 584nm

wavelength

  • Intensity of red linked to

amount of transcription from the promoter

  • No fluorescence seen in

PfadR cells – indicates no transcription from the promoter

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Expert Consultation

Why?

  • Understand urban drainage and the water industry

Who?

  • Two leading academics working with water industry

What?

  • Implications for product design

– Compatible with wastewater treatment processes – Consider GMO release issues

Framework Method Analysis Output Output

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

FOG Problems: Who’s to blame?

Reason for study

  • Develop insight from first consultations

into the webs of responsibility relating to FOG management Methodology

  • Semi-structured interviews and visits

designed to find out about differing views

  • n notions of responsibility

Framework Ethics Methods Analysis Output Ethics Ethics

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

FOG Problems: Methodology

Key Stakeholders

  • 6 Restaurant & Homeowner
  • 3 Water Industry
  • 1 Council

FOG problems at Hayton wastewater treatment facility

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

FOG Problems: Findings

Findings

  • Wide ranging practices
  • Law on side of water industry
  • Consumer ratings
  • Limitations
  • Who’s to blame?

Private company collecting used cooking oil

Complex web of practices

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Nested Manufacturing

The bacteria in the unit

The lipase from the bacteria

The unit itself

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Product Development

Framework Expert consultation Personal research Method Analysis Output 8 Concepts Pro’s and con’s Finished design Method

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

In-Sewer Concept

Reduces sewer capacity Larger scale installation that is easier to enforce and upkeep

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Grease Trap Concept

Doesn’t protect from detergents and is too BIG Already in use in restaurants and hence can modify existing units

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Perfusion Bioreactor

Too EXPENSIVE

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Vertical Stirred Tank Concept

Low oxygen diffusion

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

The Fatberglar

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Summary

Easy to maintain 30 days between refills Small enough for the kitchen sink Cheap to buy less than £100 Cheap to run • 132ml of water per day

  • bleach once a month
  • freeze dried bacteria
  • one nutrient block per

month

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Final Design

Criteria

  • Secrete low dose lipase and keratinase

enzymes continuously

CONSTITUTIVE KERATINASE (BBa_K1498000)

Constitutive promoter RBS kerUS Keratinase Terminators BBa_J23110 Bba_K1498000 Constitutive promoter RBS tliA Lipase Terminators BBa_J23110 BBa_K258006

PtetR RBS PrtD Domain Terminator RBS PrtE Domain Terminator RBS PrtF Domain Terminators BBa_R0040 B0034 K258100 B0010 B0034 K258101 B0010 B0034 K258102 B0010, B0012

CONSTITUTIVE LIPASE (BBa_K1498001) TYPE I SECRETION MODULE (BBa_K258008)

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Results: Lipase

Assay for production of fatty acids by breakdown of lipids

Empty Plasmid & Exporter Promoter, Lipase & Exporter 10 µl of cell culture supernatant on Rhodamine B plate

Proof that our lipase works and is exported

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Results: Keratinase

Assay to show production and secretion of keratinase using detection

  • f general proteases.

BBa_J23110 BBa_J23110 & Keratinase

  • S. aureus

Suspected keratinase

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Results

  • Successfully designed a system to

break down fat and hair.

  • Our design evolved based on advice

from the product design team.

  • Produced a working lipid breakdown

system.

  • Added a promoter device to the

already existing lipase biobrick.

  • Cloned Keratinase into a submission

vector as a new biobrick.

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Summary

Working lipid breakdown system Realistic delivery system

Expert Consultation Notions of Responsibility

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INTRODUCTION SOCIOBRICKS INITIAL DESIGN POLICY & PRACTICES MODELLING PRODUCT FINAL DESIGN & RESULTS SUMMARY ATTRIBUTIONS

Attributions

The University Of Sheffield Chemical And Biological Engineering

  • Prof. Catherine Biggs
  • Dr. Paul Dobson
  • Dr. Robert Falconer
  • Simon Hall
  • Dr. Henriette Jensen
  • David Wengraf
  • Prof. Phillip Wright

Civil Engineering

  • Prof. Simon Tait

Mechanical Engineering

  • Hugh Clark

Sociological Studies

  • Kate Bulpin

The School Of Clinical Dentistry

  • The Graham Stafford Research Group
  • Brenka Mccabe And Jason Heath

The Faculties Of Arts And Humanities, Engineering, Medicine, Dentistry And Health, Science And Social Science The University Of Manchester

  • Dr. Andrew Balmer

Yorkshire Water Anglian Water Northumbrian Water Sheffield City Council

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Questions