Team:Freiburg/parts/favorite parts

From 2013.igem.org

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<p> Our uniCAS Activator  
<p> Our uniCAS Activator  
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No. 2: BBa_K1150024 - uniCAS Histone Modifier
No. 2: BBa_K1150024 - uniCAS Histone Modifier
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<td> <img class="imgtxt" width="700px" src="https://static.igem.org/mediawiki/2013/archive/b/bc/20131003214618%21Freiburg2013_Plasmid_Cas9-G9a.png"> </td>
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<td> <b>Figure X: CMV:dCas9-G9a (<a id="link" href="http://parts.igem.org/Part:BBa_K1150024">BBa_K1150024</a>) </b><br>
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dCas9 was fused via a 3 amino acid linker to G9a. The resulting fusion protein was flanked by NLS sequences and tagged by a HA epitope. The CMV promoter and BGH terminator were chosen to control gene expression.
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<p>This device combines the dCas9 protein with the SET-domain of the murine histone methyltransferase G9a. dCas9 enables not only sequence specific, but also multiple targeting of any requested DNA sequence. Hence, coupling of dCas9 to the effector G9a allows for specific methylation of histone. <br>
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Such methylations are a hallmark of gene repression. One interesting fact about histone modification is the capability to spread the activity state over the surrounding chromatin via reader proteins. So the information of e.g. "repressed state" can, once specifically introduced, be propagated over a whole locus.<br><br>
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We included the G9a histone methylase to the uniCAS toolkit for specific histone methylation. With the device <a id="link" href="http://parts.igem.org/Part:BBa_K1150024">BBa_K1150024</a>, consisting of dCas9 and G9a, we were able to decrease the endogenous VEGF expression in HEK293T cells about 50%, depending on the locus targeted (Figure XXXXXX). <br><br>
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<td> <img class="imgtxt" width="700px" src="https://static.igem.org/mediawiki/2013/f/ff/G9a_pd_neu_Freubrug_2013.png"> </td>
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<td> <b>Figure X: Endogenous, stable repression by dCas9-G9a </b><br>
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Chromatin remodeling, resulting in repression of endogenous genes, is possible by fusing the histone methyltransferase G9a to dCas9. (n=3, p<0.05 is marked by asterisks)
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dCas9-G9a is our most efficient repressiv device!
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Revision as of 15:57, 4 October 2013


Our favorite BioBricks

No. 1: BBa_K1150020 - uniCAS Activator

Our uniCAS Activator

No. 2: BBa_K1150024 - uniCAS Histone Modifier

Figure X: CMV:dCas9-G9a (BBa_K1150024)
dCas9 was fused via a 3 amino acid linker to G9a. The resulting fusion protein was flanked by NLS sequences and tagged by a HA epitope. The CMV promoter and BGH terminator were chosen to control gene expression.

This device combines the dCas9 protein with the SET-domain of the murine histone methyltransferase G9a. dCas9 enables not only sequence specific, but also multiple targeting of any requested DNA sequence. Hence, coupling of dCas9 to the effector G9a allows for specific methylation of histone.
Such methylations are a hallmark of gene repression. One interesting fact about histone modification is the capability to spread the activity state over the surrounding chromatin via reader proteins. So the information of e.g. "repressed state" can, once specifically introduced, be propagated over a whole locus.

We included the G9a histone methylase to the uniCAS toolkit for specific histone methylation. With the device BBa_K1150024, consisting of dCas9 and G9a, we were able to decrease the endogenous VEGF expression in HEK293T cells about 50%, depending on the locus targeted (Figure XXXXXX).

Figure X: Endogenous, stable repression by dCas9-G9a
Chromatin remodeling, resulting in repression of endogenous genes, is possible by fusing the histone methyltransferase G9a to dCas9. (n=3, p<0.05 is marked by asterisks)
dCas9-G9a is our most efficient repressiv device!

No. 3: BBa_K1150034 - uniCAS RNAimer

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