Team:Wash U/Biological Parts
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<br><br><br><br><br><br><br><br><br><br><br>[[Image:Slide3.jpg|480px|left]] The next step in our characterization of our synthetic red light response system is to analyze the phosphorylation of ompR in ''Rhodobacter sphaeroides''. In our final system, we only want puc genes to be transcribed and expressed via the phosphorylation of ompR, not simply the puc promoter as it naturally occurs. By placing the ompR coding region downstream of the red light sensor and upstream of a terminator our modified system controls expression of the puc genes by the red light sensor in addition to the puc promoter. It should be impossible for the puc promoter to directly cause the transcription of puc genes due to the terminator, but instead, the puc genes must be activated via ompR phosphorylation. The end target of ompR transcription is the ompC promoter, located directly upstream of the puc genes. Placing GFP behind ompC tells us how often the promoter is transcribed and how often ompR is phoshorylated. | <br><br><br><br><br><br><br><br><br><br><br>[[Image:Slide3.jpg|480px|left]] The next step in our characterization of our synthetic red light response system is to analyze the phosphorylation of ompR in ''Rhodobacter sphaeroides''. In our final system, we only want puc genes to be transcribed and expressed via the phosphorylation of ompR, not simply the puc promoter as it naturally occurs. By placing the ompR coding region downstream of the red light sensor and upstream of a terminator our modified system controls expression of the puc genes by the red light sensor in addition to the puc promoter. It should be impossible for the puc promoter to directly cause the transcription of puc genes due to the terminator, but instead, the puc genes must be activated via ompR phosphorylation. The end target of ompR transcription is the ompC promoter, located directly upstream of the puc genes. Placing GFP behind ompC tells us how often the promoter is transcribed and how often ompR is phoshorylated. | ||
<br><br><br><br><br><br><br><br><br>[[Image:Slide4.jpg|480px|left]] Part three of our characterization measures the effectiveness of the red light sensor in phosphorylating ompR. This setup is the exact same as in part two except that we have added in the red light sensor. Now, ompR may be phosphorylated by the puc promoter or the red light sensor instead of solely by the puc promoter. GFP is still attached to the end product, the ompC promoter. By comparing the fluorescence of GFP in this scenario compared with the second scenario we will see how much more ompR was phosphorylated with the red light sensor that without. | <br><br><br><br><br><br><br><br><br>[[Image:Slide4.jpg|480px|left]] Part three of our characterization measures the effectiveness of the red light sensor in phosphorylating ompR. This setup is the exact same as in part two except that we have added in the red light sensor. Now, ompR may be phosphorylated by the puc promoter or the red light sensor instead of solely by the puc promoter. GFP is still attached to the end product, the ompC promoter. By comparing the fluorescence of GFP in this scenario compared with the second scenario we will see how much more ompR was phosphorylated with the red light sensor that without. | ||
- | <br><br><br><br><br><br><br><br><br><br><br><br><br>[[Image:Slide5.jpg|480px|left]] | + | <br><br><br><br><br><br><br><br><br><br><br><br><br>[[Image:Slide5.jpg|480px|left]] This is the final construct that will be our actual functioning model in ''Rhodobacter sphaeroides''. This finished product will be compared to the wild type over various intensities of light and cell culture densities can be compared to see which strain, the wild type or the |
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Revision as of 17:09, 10 July 2009