Difference between revisions of "Team:Paris Saclay"
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<p style="font-size:11pt"> The iGEM Paris-Saclay project is part of the Foundational Advance track, and aims to study the effects of DNA topology on gene expression in E.coli. The purpose is to answer to this question: ''Does bringing a strong promoter closer to a weak promoter influence the expression level of genes located downstream?'' | <p style="font-size:11pt"> The iGEM Paris-Saclay project is part of the Foundational Advance track, and aims to study the effects of DNA topology on gene expression in E.coli. The purpose is to answer to this question: ''Does bringing a strong promoter closer to a weak promoter influence the expression level of genes located downstream?'' | ||
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<p style="font-size:11pt">In the laboratory, we first focused on the tool used to visualize the interaction between both dCas9. For that purpose, we designed a biobrick in order to characterize the assembly of the split GFP. This biobrick is composed of one part of FRB / FKBP12 system fused to the other part of tripartite split-GFP system (GFP 10 / GFP 11) plus GFP 1.9 in the same plasmid. This biobrick was characterized by measuring GFP activity. Furthermore, a model was built in order to determine the optimal distance between the two dCas9 proteins for GFP to fluoresce. In order to test this model, we designed one biobrick composed of tripatrite split-GFP plus two dCas9 and another biobrick composed of the two target sequences of the dCas9 and the two sgRNA coding sequence. For this second biobrick, we wanted to test several distances (50 bp, 75 bp, 100 bp and 150 bp) between the two target sequences of the dCas9, in order to determine the best distance for tripartite split-GFP to fluorescence, regarding to the established model.</p><br> | <p style="font-size:11pt">In the laboratory, we first focused on the tool used to visualize the interaction between both dCas9. For that purpose, we designed a biobrick in order to characterize the assembly of the split GFP. This biobrick is composed of one part of FRB / FKBP12 system fused to the other part of tripartite split-GFP system (GFP 10 / GFP 11) plus GFP 1.9 in the same plasmid. This biobrick was characterized by measuring GFP activity. Furthermore, a model was built in order to determine the optimal distance between the two dCas9 proteins for GFP to fluoresce. In order to test this model, we designed one biobrick composed of tripatrite split-GFP plus two dCas9 and another biobrick composed of the two target sequences of the dCas9 and the two sgRNA coding sequence. For this second biobrick, we wanted to test several distances (50 bp, 75 bp, 100 bp and 150 bp) between the two target sequences of the dCas9, in order to determine the best distance for tripartite split-GFP to fluorescence, regarding to the established model.</p><br> | ||
Revision as of 13:39, 7 October 2016
