Kingston, Canada Queens_Canada iGEM 2011 Americas Jamboree content - - PowerPoint PPT Presentation

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Kingston, Canada Queens_Canada iGEM 2011 Americas Jamboree content - - PowerPoint PPT Presentation

Kingston, Canada Queens_Canada iGEM 2011 Americas Jamboree content the problem The Challenge Our solution Approach & Methods Future Applications Accomplishments one organism. one solution. degrade report toxins target chemical


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Queens_Canada iGEM 2011 Americas Jamboree Kingston, Canada content


 The Challenge Our solution Approach & Methods Future Applications Accomplishments

the problem

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NEMOREMEDIATION move to to contaminated site report toxins degrade target chemical Sustainable/ non-intrusive approach

  • ne organism. one solution.

why C. elegans?

  • Excellent(chemotaxis(

mechanism(

  • Mul12cellular=(mul1ple(

processes(

  • Hardy(organism(
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part design

Interchange(GPCR(genes(

*Teng et al., 2006; UCSF 2010

Chemotaxis Parts

Promoter' Effect' ODR7( A?rac1ve( STR2( A?rac1ve(( GYC7( Repulsive( GPC1( Repulsive( GPCR' Source' Ligand' ADBR2( Mouse( Phenol( Orexin( Human( Toluidine,(42Chlorobenzoic(acid,(( Naphthalene(( OX1R( Mouse( 42Chlorobenzoic(acid( MARM1( Rat( Phenol( NPYR1( Mouse( Naphthalene(

  • Conducted(preferen1al(assay(between(target((

and(control(substances(

chemotactic assay

!"#$% = #!!"!#$!!"#$%!!"!!"#$%&!!"#$%#&'! − #!!"!#$!!"#$%!!"!!"#$%"&!!"#$%#&'! #!!"!#$!!"#$% ! !

results

Part: ODR-7:ADBR2:UNC-54 (BBa_K631032)

20.2( 0( 0.2( 0.4( 0.6( 0.8( 1(

0.03( 0.05( 0.08( Chemotactic Index Concentration (g/mL)

Transgenic Control

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naphthalene degradation

Naphthalene(converted(to(salicylic(acid(

naphthalene degradation

  • 22hydroxychromene222

carboxylate(isomerase((

  • nahD(
  • Pseudomonas*pu,da(

part: BioBrick nahD

  • Characterize(in(C.*elegans((
  • Assembly(with(other(enzymes(in(C.*elegans,(

create(full(pathway(

  • BioBrick(Registry:(develop(bioremedia1on(

pathways(for(other(chassis((( (

future applications

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part improvement

  • Modify(exis1ng(eCFP(part(
  • Test(new(promoters(effec1vely((

injection reporter

  • C. elegans neurons

BioBrick Part: BBa_K6310
 GPC-1:eCFP:UNC-54:1C3 Expression in AWB neuron

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SLIDE 6

kill switch mechanism

Target(Gene(

conventional methods of BioBrick assembly

standard assembly 3-way assembly

conventional method of BioBrick assembly

1C3 1T3 1A3

novel assembly: PCR ligation

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Faculty Advisors: Dr. Chin-Sang, Dr. Greer, Dr. Ko, Dr. Bendena, Dr. Ramsay, & Dr. Young. Lab Support: Tony Papanicolaou, Jeff Boudreau and rest of Chin-Sang lab. Past QGEM Team Members: Steve Goldie, Chris Palmer, Geoff Halliday, Harry Zhou and James MacLeod. QGEM 2011: Adrian McNeely, Tony He, Stephanie Zhou, Olivia Margie, Alex Campbell, Allister Smith, David Dai, Sophie Gong, Kevin Chen, Kristina Collavino, Eni Rukaj, Faisal Bakhteyar, Eszter Gereb, Gary Wu, Wei Liu, Anujan Poologaindran. QGEM 2010/2011 Executive Team

acknowledgements

((

sponsors

1.(Ahmed(S,(Hodgkin(J.((2000)(MRT22(checkpoint(protein(is(required(for(germline(immortality(and(telomere(replica1on(in(C.*elegans.(Nature,(403:1592164(( ( 2.(Smelick(C,(Ahmed(S.((2005)(Achieving(immortality(in(the(C.*elegans(germline.(Ageing(Research(Reviews,(4(1):67282(( ( 3.(Telomere(Shortening(and(Damage((2009)((h?p://www.immortalhumans.com/telomere2shortening2and2damage/)( ( 4.(Fire(A,(Xu(S,(Montgomery(MK,(Kostas(SA,(Driver(SE,(Mello(CC.((1998)(Potent(and(specific(gene1c(interference(by(double2stranded(RNA(in(Caenorhabdi1s( elegans.(Nature,(391(6669):806211( ( 5.(Tabara(H,(Grishok(A,(Mello(CC.((1998)(RNAi(in(C.*elegans:(soaking(in(the(genome(sequence.(Science,(282(5388):43021( ( 6.(Timmons(L,(Fire(A.((1998)(Specific(interference(by(ingested(dsRNA.(Nature,(395(6705):854( ( 7.(Timmons(L.((2006)(Construc1on(of(Plasmids(for(RNA(Interference(and(In(Vitro(Transcrip1on(of(Double2Stranded(RNA.(Methods(Mol(Biol.(351:109217.( ( 8.(Simmer(F,(Tijsterman(M,(Parrish(S,(Koushika(SP,(Nonet(ML,(Fire(A,(Ahringer(J,(Plasterk(RHA.((2002)(Loss(of(the(Puta1ve(RNA2Directed(RNA(Polymerase( RRF23(Makes(C.(elegans(Hypersensi1ve(to(RNAi.(Current(Biology,(12(15):(131721319.( ( 9.(Kiyohara(H,(Torigoe(S,(Kaida(N,(Asaki(T,(Iida,(T,(Hayashi(H(and(Takizawa(N.((1994)(Cloning(and(Characteriza1on(of(a(Chromosomal(Gene(Cluster,(pah,(That( Encodes(the(Upper(Pathway(for(Phenanthrene(and(Naphthalene(U1liza1on(by(Pseudomonas(pu1da(OUS82.(Journal(of(Bacteriology,(176:243922443.(( ( Graphics'(order(of(appearance)( h?p://s.ngm.com/2009/03/canadian2oil2sands/img/candian2oil2sands2615.jpg(( h?p://en.wikipedia.org/wiki/GPCR(( h?p://ginkgobioworks.com/support/BioBrick_Assembly_Manual.pdf( h?p://www.ncbi.nlm.nih.gov/pmc/ar1cles/PMC205370/((( h?p://www2fgg.eur.nl/ch1/cellbiology/images/jansen1.jpg((

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