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        <font size="100"><h3>Bikard et al. used dCas9-ω targetting the promoter PAM rich URS J23117, BBa_K1723001, in order to regulate gene expression using gRNA (single guide RNA) guided dCas9-ω. By using our own dCas9-ω system we proved that this promoter can be activated or repressed (see <a href="https://2015.igem.org/Team:EPF_Lausanne/Results">results page</a>). Now, on the model of this promoter, we created <b>a new fully synthetic promoter</b>: PAM rich URS J23117Alt promoter, BBa_K1723005. We mutated the sequence of BBa_K1723001 between and outside the -10 and -35 sequence where the RNA polymerase binds (see <a href="https://2015.igem.org/Team:EPF_Lausanne/Results">BBa_K1723005 registry page</a> for more details)</h3></font> in order to have <b>a promoter targeted by a set of sgRNAs different</b> from the sgRNAs aiming for BBa_K1723001. The creation of this part and its experimental validation (see <a href="https://2015.igem.org/Team:EPF_Lausanne/Results">results page</a>) is very promising for us as it is the proof of the MUTABILITY of the targeted promoters. We can now imagine of mutating again the promoter to obtain others promote/sgRNAs sets creating more and more different transistors-like elements in cells. 
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<h2>PAM rich URS J23117Alt promoter</h2>
 
<h2>PAM rich URS J23117Alt promoter</h2>
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<p>This promoter was made on the same template as PAM rich URS J23117 promoter, BBa_K1723001, but was mutated to be targeted by different sgRNAs. It is also fused to a PAM rich upstream regulatory region (URS). Remember that dCas9 can bind only to PAM-flanked target sites.</p>
 
 
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<a href="http://parts.igem.org/Part:BBa_K1723005"><b>BBa_K1723005</b></a>
 
<a href="http://parts.igem.org/Part:BBa_K1723005"><b>BBa_K1723005</b></a>
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        <font size="4">Bikard et al. used dCas9-ω targeting the promoter PAM rich URS J23117, BBa_K1723001, in order to regulate gene expression [1], using gRNA (single guide RNA). By using our own dCas9-ω system we proved that this promoter can be activated or repressed (see <a href="https://2015.igem.org/Team:EPF_Lausanne/Results">results page</a>). Now, on the model of this promoter, we designed <b>a new, fully synthetic, promoter</b>: PAM rich URS J23117Alt promoter, BBa_K1723005. We mutated the sequence of BBa_K1723001 between and outside the -10 and -35 sequences where the RNA Polymerase binds (see <a href="https://2015.igem.org/Team:EPF_Lausanne/Results">BBa_K1723005 registry page</a> for more details), in order to have <b>another promoter targeted by a different set of sgRNAs</b>. The creation of this part, and its experimental validation (see <a href="https://2015.igem.org/Team:EPF_Lausanne/Results">results page</a>), is very promising for us as it is the proof of the MUTABILITY of the promoters. We can now imagine of designing other new sequences to obtain others promoter/sgRNAs sets creating more and more different transistors-like elements in cells.</font>
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        <h3>References</h3>
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        <p>[1] Bikard, D., Jiang, W., Samai, P., Hochschild, A., Zhang, F., & Marraffini, L. A. (2013). Programmable repression and activation of bacterial gene expression using an engineered CRISPR-Cas system. Nucleic acids research, 41(15), 7429-7437.</p>
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Latest revision as of 03:07, 19 September 2015

EPFL 2015 iGEM bioLogic Logic Orthogonal gRNA Implemented Circuits EPFL 2015 iGEM bioLogic Logic Orthogonal gRNA Implemented Circuits

OUR BEST NEW PART




PAM rich URS J23117Alt promoter






BBa_K1723005

Bikard et al. used dCas9-ω targeting the promoter PAM rich URS J23117, BBa_K1723001, in order to regulate gene expression [1], using gRNA (single guide RNA). By using our own dCas9-ω system we proved that this promoter can be activated or repressed (see results page). Now, on the model of this promoter, we designed a new, fully synthetic, promoter: PAM rich URS J23117Alt promoter, BBa_K1723005. We mutated the sequence of BBa_K1723001 between and outside the -10 and -35 sequences where the RNA Polymerase binds (see BBa_K1723005 registry page for more details), in order to have another promoter targeted by a different set of sgRNAs. The creation of this part, and its experimental validation (see results page), is very promising for us as it is the proof of the MUTABILITY of the promoters. We can now imagine of designing other new sequences to obtain others promoter/sgRNAs sets creating more and more different transistors-like elements in cells.

References

[1] Bikard, D., Jiang, W., Samai, P., Hochschild, A., Zhang, F., & Marraffini, L. A. (2013). Programmable repression and activation of bacterial gene expression using an engineered CRISPR-Cas system. Nucleic acids research, 41(15), 7429-7437.

EPFL 2015 iGEM bioLogic Logic Orthogonal gRNA Implemented Circuits