2
3
62% of the people we interviewed are totally ignorant about this danger 4
And you ? 5
PCBs 6
PCBs P oly C hloro B iphenyls • A family of synthetic compounds • High chemical and thermal stability 7
Utilisation of PCBs • Mostly used as insulating fluids in electrical equipment • But also in coolants, paints and cements, as stabilizing additives in flexible PVC, pesticides… 8
Production of PCBs • Produced and used during the 2 nd half of the 20 th century 400 million tons produced worldwide 9
PCB toxicity Increasing PCB concentration Water and soil contamination 10
PCB toxicity Most PCBs are : ‒ Carcinogenic Increasing PCB concentration ‒ Endocrine disruptors Dangerous for human health and the environment Water and soil contamination Ban of PCBs production and use in France in 1987 11
PCBs in the environment • Contaminated soils and waters (sediments) Paris 28% of French population : Higher exposure than PTMI (Provisional Tolerable Monthly Intake) Sites polluted by PCBs in France 12 Ref : Association Robin des Bois, from data from Ineris, www.pollutionsindustrielles.ecologie.gouv.fr
13
Are PCBs biodegradable ? First PCB degrading bacteria discovered here! Hudson river • Bacterial communities can degrade PCBs • Bacteria degrading PCBs have been isolated and studied • But none of them can totally degrade PCBs 14
2 step PCB degradation ANAEROBIC PATHWAY (e.g. Dehalobium chlorocoercia ) Highly chlorinated PCBs Lowly chlorinated PCBs Dechlorination AEROBIC PATHWAY (e.g. Burkholderia xenovorans LB 400 ) Lowly chlorinated PCBs Acetyl coenzyme A Oxidative degradation
PCB degradation pathways chosen ANAEROBIC PATHWAY (e.g. Dehalobium chlorocoercia ) tceA AEROBIC PATHWAY (e.g. Burkholderia xenovorans LB 400 ) A1 A2 orf3 A3 A4 B C
Objectives of the PCB Busters ’ project To combine the anaerobic and aerobic degradation 1 pathways in a single bacterium Choice of the chassis: E. coli -Quick growth -Dissemination control -Aerobic and anaerobic metabolisms 2 To construct a PCB sensor system 17
PCB sensor system Objective: build bacteria strain able to detect PCBs in water
PCB sensor system In a Pseudomonas pseudoalcaligenes strain: PCB PCB BphR2 BphR2 Inactive Active PCB BphR2 PbphR1 BBa_K1155001 PbphA1 Bba_K1155002
PCB sensor system PCB Blue signal BphR2 lacZ PbphR1 bphR2 Constitutive PbphA1 promoter +PCB BphR2 is active Blue signal -PCB BphR2 is inactive No signal
PCB sensor system BphR2 lacZ PbphR1 bphR2 Constitutive PbphA1 promoter +PCB BphR2 is active Blue signal -PCB BphR2 is inactive No signal
PCB degradation system Objective: build bacteria able to completely degrade PCBs.
PCB degradation system It involves two pathways: - Anaerobic: reductive dechlorination - Aerobic: oxidative degradation - No bacteria are able to perform both pathways Objective: To combine aerobic and anaerobic steps of PCB degradation in E.coli
PCB degradation system In a E. coli strain: O 2 FNR FNR Active Inactive FNR Pndh* BBa_K1155000 PnirB BBa_K1155004 PnarG BBa_K1155005 PnarK BBa_K1155006
Expression of all the PCB degradation enzymes in E. coli FNR Enzyme fnr PnarK Reductive Constitutive PnarG dechlorination promoter PnirB -O 2 FNR is active Reductive Dechlorination No +O 2 FNR is inactive No Oxidative degradation
Expression of all the PCB degradation enzymes in E. coli FNR Enzyme fnr PnarK Reductive Oxidative Pndh* Constitutive PnarG dechlorination degradation promoter PnirB -O 2 FNR is active Reductive Dechlorination No +O 2 FNR is inactive No Oxidative degradation
Expression of all the PCB degradation enzymes in E. coli FNR O 2 fnr PnarK Reductive Constitutive PnarG dechlorination promoter PnirB -O 2 FNR is active Reductive Dechlorination No +O 2 FNR is inactive No Oxidative degradation
Expression of all the PCB degradation enzymes in E. coli FNR O 2 Enzymes fnr PnarK Reductive Oxidative Pndh* Constitutive PnarG dechlorination degradation promoter PnirB -O 2 FNR is active Reductive Dechlorination No +O 2 FNR is inactive No Oxidative degradation
Testing the system Reporting gene AmilCP (Uppsala 2011 - BBa_K592009) AmilCP
Testing the system Reporting gene AmilCP (Uppsala 2011 - BBa_K592009) AmilCP Term RBS
Testing the system FNR fnr RBS AmilCP Pndh* Term Constitutive promoter -O 2 FNR is active Pndh* repressed No +O 2 FNR is inactive Pndh* no repressed Violet signal
Pndh* - amilCP expression ANAEROBIC CONDITIONS AEROBIC CONDITIONS
Summary Work done: Work in progress: • three FNR activated • Finish the sensor system promoters • Characterize NarG, NarK • one FNR repressed promoter and NirB promoters • Improved a reporter system • Finish the construction of the degradation system • Characterize Pdnh* promoter by using RBS_AmilCP_Term construction
Modeling The O 2 switching system 34
HSIM Patrick Amar and Loïc Paulevé. HSIM: an hybrid stochastic simulation system for 35 systems biology, Electronic Notes in Theoretical Computer Science
HSIM reactions A → B + C [P] 36
HSIM reactions A → B + C [P] A + B → C + D [P] 37
Random walk simulation scheme A 38
Random walk simulation scheme A B 39
Random walk simulation scheme A B A+B → C+D REACTION PROBABILITY P ? 40
Random walk simulation scheme C D A+B → C+D 41
Modeling our oxygen sensor FNR Bound O 2 FNR FNR Biological system Computer model 42
Modeling our oxygen sensor FNR Bound O 2 FNR FNR Biological system Computer model FNR FNR + O 2 43
Modeling our oxygen sensor FNR Bound O 2 FNR FNR Biological system Computer model FNR FNR + O 2 Bound FNR + FNR 44 44
Modeling our oxygen sensor FNR Bound O 2 FNR FNR Biological system Computer model FNR FNR + O 2 Bound FNR + FNR + Bound FNR FNR 45 45 45
Modeling our oxygen sensor FNR Bound O 2 FNR FNR Biological system Computer model FNR FNR + O 2 Bound FNR + FNR + Bound FNR FNR Bound Bound FNR + FNR
Results Green reporter (lack of oxygen) Red reporter (oxygen present) FNR bound to promoter sites 47
A lighter HSIM simulator, wiki-integrated ! 48
A lighter HSIM simulator, wiki-integrated ! 49
A lighter HSIM simulator, wiki-integrated ! 50
A lighter HSIM simulator, wiki-integrated ! 51
A lighter HSIM simulator, wiki-integrated ! 52
A lighter HSIM simulator, wiki-integrated ! 53
A lighter HSIM simulator, wiki-integrated ! 54
A lighter HSIM simulator, wiki-integrated ! 55
A lighter HSIM simulator, wiki-integrated ! 56
A lighter HSIM simulator, wiki-integrated ! 57
A lighter HSIM simulator, wiki-integrated ! 58
A lighter HSIM simulator, wiki-integrated ! 59
A lighter HSIM simulator, wiki-integrated ! 60
Human Practices Perception of our project by the public Safety Open Source 62
Perception of our project by the public Yes What do you think about No our PCBbuster project ? Have you ever heard In favor about PCBs ? Against No opinion 63
Perception of our project by the public Safety questions Intellectual Property 64
PS Bacterial Filter Gemote system Sensor detection Anaerobic and aerobic combination 65
Open Source Reflection Meetings : - Comparison free software/synthetic biology - Bioterrorism history - Democratization of science - Bioeconomy - Biohacking Interview : Ellen Jorgensen « I think that people are curious » « DNA is everywhere » « Synthetic biology is very powerful! » 66
Open Source Reflection PS PCR : open source thermo cycler 67
PS-PCR : heating/cooling 68
PS-PCR : heating/cooling Peltier element (3.78 € ) 69
PS-PCR : heating/cooling Peltier element (3.78 € ) Heat sink 70
PS-PCR : sample holder Electrolab.fr 71
PS-PCR : thermal block 72
PS-PCR : control electronics 73
PS-PCR : computer software Open source control software, in collaboration with Pierre-Yves 74
PS-PCR : minimal cost 75
Test PS-PCR 15µL 76
Conclusion PbphR1 PbphA1
Thank you ! Super advisors Claire Nguyên Solenne Sylvie Jean-Luc Philippe
USING SPORT FOR PREVENTING VIOLENCE AND INTERNET ADDICTION Submitting author:
Examining Feedback Surveys from NCI Interviews in Florida What was the
Robert E Wiltbank, Ph.D. wiltbank@willamette.edu VERY Active Angels
Methodology Interviewed 94 leaders from NGOs, contractors, government
Nave Organ Project Update 2018 Annual Meeting 2017 Committee Activity
Assignment #1: Needfinding Introduction Education for social mobility Cathy
Pe mbina Hills Public Sc hools Communic ations plan Pub lishe d F e b . 11,
Presentation to SMIG 28 th February 2013 People People are our most valued
Evidence- Based Planning Session 2 March 22, 2016 Review Clusters you
People Either Drain Or Replenish Others. People Problems Or Power People Over
7/7/2020 Operation of Activated Sludge Denitrification and Total Nitrogen
Meeting 4 Physical Fitness Lead: Bill Kraus Members: Kirk Erickson, Kathy
For youngsters and their families Dr Alistair Lumb, Diabetologist Dr Taffy
How Much is E N N OUGH? OUGH? How Much is E Optimizing Nitrification and
G.D GOENKA PUBLIC SCHOOL, SARITA VIHAR Yoga Yoga & & Aer Aerobics
W3: Individual Fitness Fitness Exercise Testing Pyschology Exercise
2006) Sex Roles, Vol. 54, Nos. 3/4, February 2006 ( C DOI:
MWEA Annual Conference June 26, 2012 ORP Improves Operational Efficiency
Post-Polio Syndrome and Exercise Julie Simpson, PT Neuro and Cancer Rehab
Gr Groundwater r Remedia iatio ion n an and d Dual al-Bi Biofi ofilm
Regional Conference Presentation Planning Considerations Planning
Sustainable Water Treatment And Reuse Peter L.Timpany 2017.05 Registered