Difference between revisions of "Team:Bielefeld-CeBiTec"

 
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iGEM Bielefeld-CeBiTec 2017
 
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<h2> Expanding the Genetic Code </h2>
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<article> We are exploring the application of unnatural base pairs as an expansion of the genetic code. To prevent loss of unnatural base pairs during replication, we will utilize several systems including CRISPR/Cas9. The expanded genetic code allows for the ribosomal incorporation of multiple non-canonical amino acids (ncAAs) into peptides. With their broad chemical and functional diversity, ncAAs provide a variety of promising applications including protein labeling, photocaging, structure analysis, and specific protein interactions. Therefore, our innovative toolkit for the translational incorporation of ncAAs in E. coli is a valuable contribution to iGEM. Directed evolution of tRNA/aminoacyl-tRNA synthetase pairs enables the site-specific incorporation of ncAAs into peptides. This approach results in an optimal orthogonality to the autologous translation apparatus and a high flexibility concerning the incorporation of multiple ncAAs. As proof of concept, we are developing a rapid test for prions and a new chromatography method for mild protein elution.
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For more information visit our <b><a href="https://www.researchgate.net/publication/318084694_Expansion_of_the_genetic_code_for_the_translational_incorporation_of_non-canonical_amino_acids">project poster</a></b> on reasearch gate
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<h2>Achievements</h2>
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The following list sums up all of our achievements and refers to pages of this wiki containing detailed information. Next to this we recorded the <b><a href="https://youtu.be/1f5vwcrvUac">original presentation</a></b> of our project in Boston at the Giant Jamboree, which sums up and explains the project in less than 20 minutes:
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<h3>Awards</h3>
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<ul style="margin-left: 5%;">
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<li>Best Foundational Advance Project</li>
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<li>Best New Basic Part</li>
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</ul>
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<h3>Nominations</h3>
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<ul style="margin-left: 5%;">
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<li>Best Software</li>
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<li>Best Hardware</li>
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<li>Best New Composite Part</li>
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<li>Best Part Collection</li>
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</ul>
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<b>Establishment of two orthogonal methods for the detection of unnatural base pairs in a target sequence via Oxford Nanopore sequencing and an enzyme based detection method</b>
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<b>We established two orthogonal methods for the detection of unnatural base pairs in a target sequence: an <a target="_blank" href="https://2017.igem.org/Team:Bielefeld-CeBiTec/Software">Oxford Nanopore sequencing</a> application and an enzyme based detection method</b>
 
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<b>Development of a software suite for these orthogonal methods </b>
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<b>Development of a <a target="_blank" href="https://2017.igem.org/Team:Bielefeld-CeBiTec/Software">software</a> suite for these orthogonal methods </b>
 
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<b>Integration and characterization of  the nucleotide transporter PtNTT2 from <i>P.tricornutum</i> in <i>E.coli</i> for the uptake of  unnatural nucleoside triphosphates</b>
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<b>Integration and characterization of  the <a target="_blank" href="https://2017.igem.org/Team:Bielefeld-CeBiTec/Results/unnatural_base_pair/uptake_and_biosynthesis">nucleotide transporter PtNTT2</a> from <i>P.tricornutum</i> in <i>E.coli</i> for the uptake of  unnatural nucleoside triphosphates</b>
 
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<b>Proof that certain Taq-polymerases can efficiently incorporate unnatural nucleotides </b>
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<b>Confirmation that certain Taq DNA polymerases can efficiently <a target="_blank" href="https://2017.igem.org/Team:Bielefeld-CeBiTec/Results/unnatural_base_pair/preservation_system">incorporate unnatural nucleotides</a> </b>
 
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<b>Construction of a toolkit consisting of five aminoacyl-tRNA synthetases for incorporation of non-canonical amino acids</b>
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<b>Construction of a <a target="_blank" href="https://2017.igem.org/Team:Bielefeld-CeBiTec/Results/toolbox">toolbox</a> consisting of five aminoacyl-tRNA synthetases for incorporation of non-canonical amino acids</b>
 
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<b>Development of a photoswitchable lycopene pathway</b>
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<b><a target="_blank" href="https://2017.igem.org/Team:Bielefeld-CeBiTec/Results/toolbox/labeling">Colocalization</a> of the RuBisCo and and subcellular compartment (carboxysome) using a fluorescent amino acid</b>
 
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<b>Design and chemical synthesis of a novel, fully synthetic amino acid based on cyanonitrobenzothiazol and asparagine and proof of its functionality</b>
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<b>Development of a <a target="_blank" href="https://2017.igem.org/Team:Bielefeld-CeBiTec/Results/toolbox/photoswitching">photoswitchable lycopene pathway</a></b>
 
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<b>Library development with several thousand sequences for selecting aminoacyl-tRNA synthetases</b>
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<b><a target="_blank" href="https://2017.igem.org/Team:Bielefeld-CeBiTec/Model">Modeling</a> more than ten new aaRS sequences</b>
 
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<b>Construction of positive and negative selection plasmids for the evolution of new synthetases for non-canonical amino acids</b>
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<b>Writing a <a target="_blank" href="https://static.igem.org/mediawiki/2017/1/1e/T--Bielefeld-CeBiTec--DKE_Biosafety_Report.pdf">biosafety report</a> entitled “Auxotrophy to Xeno-DNA: A Comprehensive Exploration of Combinatorial Mechanisms for a High-Fidelity Biosafety System” </b>
 
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<b>Writing the <a target="_blank" href="https://static.igem.org/mediawiki/2017/1/18/T--Bielefeld-CeBiTec--CMZ-ChImp.pdf">ChImp Report</a> on “Chances and Implications of an Expanded Genetic Code”</b>
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Latest revision as of 15:18, 16 December 2017

Achievements

The following list sums up all of our achievements and refers to pages of this wiki containing detailed information. Next to this we recorded the original presentation of our project in Boston at the Giant Jamboree, which sums up and explains the project in less than 20 minutes:

Awards

  • Best Foundational Advance Project
  • Best New Basic Part

Nominations

  • Best Software
  • Best Hardware
  • Best New Composite Part
  • Best Part Collection


We established two orthogonal methods for the detection of unnatural base pairs in a target sequence: an Oxford Nanopore sequencing application and an enzyme based detection method



Development of a software suite for these orthogonal methods


Integration and characterization of the nucleotide transporter PtNTT2 from P.tricornutum in E.coli for the uptake of unnatural nucleoside triphosphates



Construction of a toolbox consisting of five aminoacyl-tRNA synthetases for incorporation of non-canonical amino acids



Colocalization of the RuBisCo and and subcellular compartment (carboxysome) using a fluorescent amino acid


Design, chemical synthesis and proof of functionality of a novel, fully synthetic amino acid based on cyanonitrobenzothiazol and asparagine



Modeling more than ten new aaRS sequences



Library development with several hundred thousand sequences for selecting aminoacyl-tRNA synthetases


Construction of positive and negative selection plasmids for the evolution of new synthetases for non-canonical amino acids



Improvement of an aminoacyl-tRNA synthetase test-system by introducing a FRET-system and development of a ranking system



Construction of an LED panel for irradiating 96-well microtiter plates, which can be used to manipulate non-canonical amino acids and for other applications



Development of an Android App to control the LED panel with your smartphone via Bluetooth


Writing a biosafety report entitled “Auxotrophy to Xeno-DNA: A Comprehensive Exploration of Combinatorial Mechanisms for a High-Fidelity Biosafety System”



Writing the ChImp Report on “Chances and Implications of an Expanded Genetic Code”