Team:Freiburg/Highlights

From 2013.igem.org

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<div id="h1">Highlights
<div id="h1">Highlights
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<div id="h2"> <p></p><b>In the last months we were able... </b><p></p> </div>  
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<div id="h2"><p></p> In the last months we were able... <p></p></div>  
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We started with mutating the nickase sites in the Cas9 protein and could generate a novel type of DNA binding protein that is relying on <b> protein-RNA-DNA </b> interaction.
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We started by mutating the nickase sites in the Cas9 protein and generated a novel type of DNA binding protein that is relying on <b> protein-RNA-DNA </b> interaction.
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By fusing <b>effector domains</b> to Cas9 gives rise to new properties. The <b>activation domain VP16</b> was able to activate transcription of genes. The fusion of the <b>transcriptional repressor domain KRAB</b> led to decreasing gene expression. Specific <b>chromatin modification</b> was achieved by fusing a histone methyl transferase G9a to dCas9. With this protein we are able to specifically repress endogenous gene expression. </p> <p> </p> <p>
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By fusing <b>effector domains</b> to Cas9 we added new properties to the protein. The <b>activation domain VP16</b> is able to activate transcription of genes. The fusion of the <b>transcriptional repressor domain KRAB</b> leads to synthetic repressor of gene expression. Specific <b>chromatin modification</b> was achieved by fusing a histone methyl transferase G9a to dCas9. With this protein we are able to specifically repress endogenous gene expression. </p> <p> </p> <p>
   
   
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Utilizing the light receptor system of higher plants we were able to dimerize the dCAS9 with the effector domain on light stimulus and in consequence activate the system.
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We were able to induce our system on light stimulus. This was possible by using photorecetors of higher plants.
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By building a plasmid containing the <b>necessary RNAs</b> and <b>insertion sites</b> for targeting we achieved a modular, BioBrick compatible system for <b>multiple DNA targeting: The RNAimer.</b>
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By building a plasmid containing the necessary<b> RNAs</b> and <b>insertion sites</b> for targeting we created a modular, BioBrick compatible system for <b>multiple DNA targeting: The RNAimer.</b>
Using our RNAimer plasmid it is easy to combine several target sequences on one plasmid using the BioBrick standard.
Using our RNAimer plasmid it is easy to combine several target sequences on one plasmid using the BioBrick standard.
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We developed an ELISA based approach, that is able to give a quantitative impression of binding strength of dCAS9 to a given target. We called this binding assay uniBAss. It is a powerful tool for the characterization of the interaction between the modified Cas9 and the locus specific RNA.
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We developed an ELISA based method. With this method we can quantify the binding efficiency of our proteins. We called this binding assay uniBAss. It is a powerful tool for the characterization of the interaction between the modified Cas9 and the locus specific RNA.
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To sum it all uo able to establish a new modularized tool kit for modulating gene expression: The uniCAS Toolkit! </p>
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In summary, we established a new modularized tool kit for modulating gene expression: The uniCAS Toolkit! </p>
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Revision as of 19:13, 28 September 2013

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