Tokyo/Works/Assay
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__NOTOC__ | __NOTOC__ | ||
| - | <br>[[Tokyo/Works|Works top]] | + | <br>[[Tokyo/Works|Works top]] 0.[[Tokyo/Works/Hybrid promoter|Hybrid promoter]] 1.[[Tokyo/Works/Formulation |Formulation]] 2.[[Tokyo/Works/Assay |Assay1]] 3.[[Tokyo/Works/Simulation |Simulation]] 4.[[Tokyo/Works/Assay2 |Assay2]] 5.[[Tokyo/Works/Future works |Future works]] |
<br><br> | <br><br> | ||
| - | [[Tokyo/Hill function fitting | | + | [[Tokyo/Hill function fitting |Objective of this assay]] [[Tokyo/AHL assay|Effect of AHL]] [[Tokyo/IPTG assay|Effect of LacI-IPTG]] [[Tokyo/Preliminary assays |Preliminary assays]] |
| - | ==[[Tokyo/Hill function fitting | | + | ==[[Tokyo/Hill function fitting |Objective of this assay (Hill function fitting) ]] == |
<!--新しく作ったハイブリッドプロモーターに関してはデータがないので，実験によりパラメータを求める． | <!--新しく作ったハイブリッドプロモーターに関してはデータがないので，実験によりパラメータを求める． | ||
ヒル関数をfittingさせることにより，n2,n3,K2,K3が求まる.--> | ヒル関数をfittingさせることにより，n2,n3,K2,K3が求まる.--> | ||
We have obtained data for our newly devised hybrid promoter. By Hill function fitting, we have determined '''n2, n3, K2, and K3'''. | We have obtained data for our newly devised hybrid promoter. By Hill function fitting, we have determined '''n2, n3, K2, and K3'''. | ||
| - | <br>[[Image: | + | <br>[[Image:siki1.JPG|300px|none|]] |
| - | <br>[[Image:expression6-10-2.JPG|320px | + | <br>[[Image:expression6-10-2.JPG|320px|left|]][[Image:siki2.JPG|400px|none|]] |
<br> | <br> | ||
<br> | <br> | ||
| - | ==== | + | ==== ==> see more [[Tokyo/Hill function fitting|details]]==== |
<!--== Assays for hybrid promoter == | <!--== Assays for hybrid promoter == | ||
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== [[Tokyo/AHL assay|Effect of AHL]] == | == [[Tokyo/AHL assay|Effect of AHL]] == | ||
| - | <br>[[Image:hill AHL.JPG|thumb| | + | <br>[[Image:hill AHL.JPG|thumb|230px|'''Fig. 1 Effect of AHL on hybrid promoter''' Enhanced fluorescence intensity represents up-regulation of LuxR via AHL.|left]] |
| - | ''' | + | '''Objective''' |
| - | + | By testing how AHL activates lux-lac hybrid promoter, parameters(n2,K2) should be determined. | |
| - | + | <br><br> | |
'''Result & Conclusion''' | '''Result & Conclusion''' | ||
| - | <br>GFP fluorescence increased in AHL-dose dependent manner, indicating that the hybrid promoter is enhanced by LuxR via AHL. Taken together with the graph shown in | + | <br>GFP fluorescence increased in AHL-dose dependent manner, indicating that the hybrid promoter is enhanced by LuxR via AHL. Taken together with the graph shown in Fig.1 and formulation of Hill function fitting described above , the characteristics of the hybrid promoter expressed in Hill function, such as (n2,K2), are determined. |
<br> | <br> | ||
| - | ==== | + | ==== ==> see more [[Tokyo/AHL assay|details]]==== |
| + | <br> | ||
| + | <br> | ||
| + | <br> | ||
| - | == [[Tokyo/IPTG assay| | + | == [[Tokyo/IPTG assay|Effect of LacI: IPTG assay]] == |
| - | [[Image:hill IPTG.JPG|thumb| | + | [[Image:hill IPTG.JPG|thumb|215px|'''Fig. 2 Effect of LacI on hybrid promoter''' Enhanced fluorescence intensity represents down-regulation of LacI repression via IPTG.|left]] |
| - | ''' | + | '''Objective''' |
| - | By testing how LacI represses lux-lac hybrid promoter, parameters(n3,K3) should be decided. In order to adjust effective concentration of LacI, IPTG was added. | + | By testing how LacI represses the lux-lac hybrid promoter, parameters(n3,K3) should be decided. In order to adjust effective concentration of LacI, IPTG was added. |
<br><br> | <br><br> | ||
'''Result & Conclusion''' | '''Result & Conclusion''' | ||
| - | GFP fluorescence intensity was enhanced in IPTG-dose dependent manner. It indicates that the LacI repression was getting weak by adding IPTG. Taken together with the graph shown in Fig. 2 and the formulation of Hill function fitting described above, the characteristics of the hybrid promoter expressed in Hill function was determined. | + | GFP fluorescence intensity was enhanced in an IPTG-dose dependent manner. It indicates that the LacI repression was getting weak by adding IPTG. Taken together with the graph shown in Fig. 2 and the formulation of Hill function fitting described above, the characteristics of the hybrid promoter expressed in Hill function was determined. |
| - | ==== | + | ==== ==> see more [[Tokyo/IPTG assay|details]]==== |
| + | <br> | ||
| + | <br> | ||
<br> | <br> | ||
<br> | <br> | ||
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We conducted preliminary assays as practice experiments and check of materials and methods. | We conducted preliminary assays as practice experiments and check of materials and methods. | ||
<br> | <br> | ||
| - | ==== | + | ==== ==> See more [[Tokyo/Preliminary assays |details]]==== |
== == | == == | ||
[[Tokyo/Works/Formulation |Before(Formulation)]] << [[Tokyo/Works/Assay |Assay1]] >> [[Tokyo/Hill function fitting|Assay1 contents]] >>>> [[Tokyo/Works/Simulation |Simulation]] | [[Tokyo/Works/Formulation |Before(Formulation)]] << [[Tokyo/Works/Assay |Assay1]] >> [[Tokyo/Hill function fitting|Assay1 contents]] >>>> [[Tokyo/Works/Simulation |Simulation]] | ||
Latest revision as of 04:53, 27 October 2007
Works top 0.Hybrid promoter 1.Formulation 2.Assay1 3.Simulation 4.Assay2 5.Future works
Objective of this assay Effect of AHL Effect of LacI-IPTG Preliminary assays
Objective of this assay (Hill function fitting)
We have obtained data for our newly devised hybrid promoter. By Hill function fitting, we have determined n2, n3, K2, and K3.
==> see more details
Effect of AHL
Objective
By testing how AHL activates lux-lac hybrid promoter, parameters(n2,K2) should be determined.
Result & Conclusion
GFP fluorescence increased in AHL-dose dependent manner, indicating that the hybrid promoter is enhanced by LuxR via AHL. Taken together with the graph shown in Fig.1 and formulation of Hill function fitting described above , the characteristics of the hybrid promoter expressed in Hill function, such as (n2,K2), are determined.
==> see more details
Effect of LacI: IPTG assay
Objective
By testing how LacI represses the lux-lac hybrid promoter, parameters(n3,K3) should be decided. In order to adjust effective concentration of LacI, IPTG was added.
Result & Conclusion
GFP fluorescence intensity was enhanced in an IPTG-dose dependent manner. It indicates that the LacI repression was getting weak by adding IPTG. Taken together with the graph shown in Fig. 2 and the formulation of Hill function fitting described above, the characteristics of the hybrid promoter expressed in Hill function was determined.
==> see more details
Appendix: Preliminary assays
We conducted preliminary assays as practice experiments and check of materials and methods.
==> See more details
Before(Formulation) << Assay1 >> Assay1 contents >>>> Simulation
