Difference between revisions of "Team:William and Mary/Description"
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<p>In addition to characterizing the noise of promoters in iGEM, we contributed additional tools for manipulating their expression. CRISPR/Cas9 has been used extensively in synthetic biology, both inside and out of iGEM. In particular, new functionalizations of the Cas9 protein that remove its catalytic nuclease domain while retaining its DNA-binding activity have allowed for novel methods in molecular biology. This catalytically inactive variant of Cas9, known as dCas9, can be used to repress gene expression by targeting the promoter region of a gene of interest. This repression is mediated by the CRISPR/Cas9 complex binding to the promoter region and can block RNAP binding or prevent transcriptional elongation. All of our gRNAs prevent RNAP binding and initiation of transcription.</p></div></p> | <p>In addition to characterizing the noise of promoters in iGEM, we contributed additional tools for manipulating their expression. CRISPR/Cas9 has been used extensively in synthetic biology, both inside and out of iGEM. In particular, new functionalizations of the Cas9 protein that remove its catalytic nuclease domain while retaining its DNA-binding activity have allowed for novel methods in molecular biology. This catalytically inactive variant of Cas9, known as dCas9, can be used to repress gene expression by targeting the promoter region of a gene of interest. This repression is mediated by the CRISPR/Cas9 complex binding to the promoter region and can block RNAP binding or prevent transcriptional elongation. All of our gRNAs prevent RNAP binding and initiation of transcription.</p></div></p> | ||
+ | <div><p>We created a codon-optimized dCas9 variant for expression in E. coli that, when used with an appropriate gRNA, reduced observed gene expression by 97%. In addition to making both a dCas9 coding region (BBa_K1795000), and a dCas9 operon for constitutive expression (BBa_K1795001), we also contribute 12 gRNAs that target commonly used promoters in iGEM, and a scrambled gRNA to be used as a negative control.</p> | ||
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+ | <p>We believe that our contributions to iGEM 2015 will not only open the door for more noise-related iGEM projects in the future, but will allow for more sophisticated synthetic gene regulatory network design both through the additional promoter characterization we have performed and the availability of new negative regulators of gene expression.</p> | ||
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Revision as of 11:53, 18 September 2015
Integrator Suites
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Antibiotic Operons
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dCas9s
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gRNAs
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XFPs Under Various Promoters
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G^2
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