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Biofilm Killer BIOSCIENCES LYON iGEM TEAM 2012 Context iGEM - PowerPoint PPT Presentation

Biofilm Killer BIOSCIENCES LYON iGEM TEAM 2012 Context iGEM Synthetic Biology for Innovative Solutions competition Context Industrial biofilm issues Oil industry : Osli, Schlumberger Cosmetics: Gatefoss Enzyme-based cleaning solutions:


  1. Biofilm Killer BIOSCIENCES LYON iGEM TEAM 2012

  2. Context iGEM Synthetic Biology for Innovative Solutions competition

  3. Context Industrial biofilm issues Oil industry : Osli, Schlumberger Cosmetics: Gatefossé Enzyme-based cleaning solutions: Realco

  4. Context � Industry / Manufacturing Unwanted biofilms Final Product Quality Cleaning Costs Use of Chemicals

  5. Context � Existing solutions Mechanical Chemical Enzymes Use of Chemicals � Impact on WWTP efficiency � Impact on Ecosystems � Registry, Evaluation, Authorisation and restriction of Chemicals (REACh)

  6. Context Objective: Use synthetic biology to reduce the quantity of chemicals used in cleaning processes

  7. Bacillus subtilis « TORPEDO » Context Project Houry, Gohar, Deschamps, Tischenko, Aymerich, Gruss & Briandet , Proc Natl Acad Sci U S A, 2012.

  8. Context Project KILL constitutive Option Option STICK COAT

  9. Context Project KILL Destroy biofilms

  10. Context Project WEAPON TARGET Staphylococcus strains Biofilm Killer : Bacillix

  11. Context Project KILL Constitutive module Lysostaphin Dispersin

  12. Biocide and scattering agents secreted inside the biofilm and not above, thanks to the swimming capacity of Bacillus .

  13. MECHANISM OF ACTION OF LYSOSTAPHIN Context Project KILL constitutive 2: Lysostaphin Binding and lysis 1: Lysostaphin production Bacillus subtilis S.aureus Lysostaphin

  14. MECHANISM OF ACTION OF DISPERSIN Context Project KILL constitutive 1: Dispersin production Bacillus subtilis 2: Scattering Effect Dispersin S.aureus

  15. Experimental Context Project work KILL constitutive Lysostaphin Dispersin 2 mechanisms of action Complementary action

  16. LYSOSTAPHIN BioBrick™ Experimental Context Project work BBa_K802000 KILL constitutive RBSb Lyso E X S P pBKL25 Er R Lysostaphin gene from Staphylococcus simulans

  17. LYSOSTAPHIN BioBrick™ Experimental Context Project work BBa_K802000 KILL constitutive 24 h Staphylococcus biofilm Upon contact with Bacillus culture for 24 h Observation of Staphylococcus biofilm with Confocal Laser Scanning Microscope

  18. LYSOSTAPHIN BioBrick™ Experimental Context Project work BBa_K802000 KILL CONTROL LYSOSTAPHIN constitutive •Biofilm S. aureus •Biofilm S. aureus •Treated with B. subtillis /empty shuttle plasmid •Treated with B. subtillis /shuttle plasmid + BioBrick™ 30 µm � BIOCIDE EFFECT OF LYSOSTAPHIN

  19. DISPERSIN BioBrick™ Experimental Context Project work BBa_K802001 KILL constitutive Disp RBSb E X S P pBKH26 Er R Dispersin gene from Aggregatibacter actinomycetemcomittans

  20. DISPERSIN BioBrick™ Experimental Context Project work BBa_K802001 KILL constitutive 24 h Staphylococcus biofilm Upon contact with Bacillus culture for 24 h Additional washing step : remove Bacillus culture and add growth medium Observation of Staphylococcus biofilm with Confocal Laser Scanning Microscope BEFORE and AFTER Washing step

  21. DISPERSIN BioBrick™ Experimental Context Project BBa_K802001 work KILL BEFORE washing constitutive DISPERSIN CONTROL •Biofilm S. aureus •Biofilm S. aureus •Treated with B. subtillis /empty shuttle plasmid •Treated with B. subtillis /shuttle plasmid + BioBrick™

  22. DISPERSIN BioBrick™ Experimental Context Project BBa_K802001 work KILL AFTER washing constitutive DISPERSIN CONTROL •Biofilm S. aureus •Biofilm S. aureus •Treated with B. subtillis /empty shuttle plasmid •Treated with B. subtillis /shuttle plasmid + BioBrick™ � SCATTERING EFFECT OF DISPERSIN

  23. Experimental Context Project work KILL constitutive LYSOSTAPHIN + DISPERSIN

  24. Experimental LYSOSTAPHIN + DISPERSIN Context Project work KILL BEFORE washing constitutive LYSOSTAPHIN + DISPERSIN CONTROL

  25. Experimental KILL LYSOSTAPHIN + DISPERSIN Context Project work constitutive AFTER washing CONTROL LYSOSTAPHIN + DISPERSIN LYSOSTAPHIN 30 µm DISPERSIN � BOTH : BETTER RESULT, CLEANER SURFACE

  26. Experimental Context Project work KILL constitutive Option Option COAT STICK Toggle switch

  27. Experimental SURFACTIN generator + BIOFILM repressor + regulator / BBa_K802009 Context Project work Avoid new biofilm Option Option COAT STICK formation P xyl RBSb RBSb RBSb sfp abrB lacI

  28. Experimental SURFACTIN generator + BIOFILM repressor + regulator / BBa_K802009 Context Project work P xyl Option Option RBSb RBSb RBSb sfp abrB lacI COAT STICK oil Emulsion VORTEX Filtered Filtered S S Bacillus culture Observation 24 h later

  29. Experimental SURFACTIN generator + BIOFILM repressor + regulator / BBa_K802009 Context Project work P xyl Option Option RBSb RBSb RBSb sfp abrB lacI COAT STICK Positive control: Supernatant Supernatant LB + SDS BBa_K802009 empty plasmid � Surfactin production

  30. Experimental SURFACTIN generator + BIOFILM repressor + regulator / BBa_K802009 Context Project work P xyl Option Option RBSb RBSb RBSb sfp abrB lacI COAT STICK ∆ ∆ abrB ∆ ∆ ∆ ∆ ∆ ∆ abrB/ BBa_K802009 WILD TYPE Biofilm No biofilm No biofilm � abrB is functional in our part

  31. Experimental Context Project Modelling work

  32. Experimental Context Project Modelling work What do we need for a model? Phenomenon Option Option STICK COAT

  33. Experimental Context Project Modelling work Formal system

  34. Experimental Context Project Modelling work Objectives

  35. Experimental Context Project Modelling work Data and Knowledge

  36. Experimental Context Project Modelling work P xyl sfp sfp abrB abrB lacI lacI XYLOSE xylR xylR P lac

  37. Experimental Context Project Modelling work COAT COAT Surfactant Biofilm (planktonic cells) regulator repressor (planktonic cells) P xyl RNA sfp abrB lacI pol + XYLOSE

  38. Experimental Context Project Modelling work P xyl sfp abrB lacI IPTG xylR P lac

  39. Experimental Context Project Modelling work No surfactant, no biofilm repressor P xyl sfp abrB lacI IPTG + RNA STICK STICK xylR pol (biofilm) (biofilm) P lac

  40. Experimental Context Project Modelling work Modelling objective: Predict the “Biofilm Killer” behavior depending on the inducers’ concentrations

  41. Experimental Context Project Modelling work Physiological Physiological response response Stick Coat Coat Stick IPTG concentration Xylose concentration

  42. Experimental Context Project Modelling work KILL constitutive IPTG induction Xylose induction Option Option STICK COAT

  43. Experimental Context Project Modelling Industrialisation work Clean surfaces Health and food industry Protect surfaces Poultry and animal farming , Chemical industry Oil industry COAT STICK Protective biofilm

  44. Experimental Human Context Project Modelling Industrialisation work Practices INDUSTRIALIST

  45. Experimental Human Context Project Modelling Industrialisation work Practices Which economic system for synthetic biology? E. OSTROM J. STIZGLITZ 2009 Nobel Prize 2001 Nobel Prize

  46. Experimental Human Context Project Modelling Industrialisation work Practices From the Commons to the open source Open source Commons, open source forerunner

  47. Experimental Human Context Project Modelling Industrialisation work Practices What is a commons ? Private sector Public sector Market State Commons Community Members « a jointly owned legal set of rights » Organised by its members

  48. Experimental Human Context Project Modelling Industrialisation work Practices Which economic system for synthetic biology ? A commons as a solution ? Do you think BioBricks TM SHOULD be patentable ? YES NO UNDECIDED

  49. Experimental Human Context Project Modelling Industrialisation work Practices Why not define together the rules of a synthetic biology commons for iGEM ?

  50. Main Achievements � A promising solution to control biofilms on closed surfaces � A proposition for an Economic System for synthetic biology (Collaboration with UBC) � Characterization of new parts : � LYSOSTAPHIN BioBrick™ /BBa_K802000 � DISPERSIN BioBrick™/BBa_K802001 � SURFACTIN + BIOFILM / BBa_K802009 � 2 shuttle plasmids BBa_K802003 and BBa_K802004

  51. Acknowledgments TEACHING FACILITIES INSA And all scientific, technical and administrative staff from : •Department of Biosciences of INSA •MAP and BF2I laboratories •MICALIS INRA laboratory And our sponsors :

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