Team:EPF-Lausanne/Tools/MITOMI

From 2011.igem.org

(Difference between revisions)
(MITOMI scans explanation)
(MITOMI scans explanation)
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[[File:EPFL2011_MITOMIchip_beforewash_illustration.png|300px| After incubation: GFP fluorescence ]]
[[File:EPFL2011_MITOMIchip_beforewash_illustration.png|300px| After incubation: GFP fluorescence ]]
[[File:EPFL2011_MITOMIchip_beforewashDNA_illustration.png|300px| After incubation: CY5 fluorescence ]]
[[File:EPFL2011_MITOMIchip_beforewashDNA_illustration.png|300px| After incubation: CY5 fluorescence ]]
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Green fluorescence comes from the GFP-tag fused to the protein or from the incorporated Green Lysine.
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Red fluorescence is due to the CY5 labeling of the DNA.
    
    
* Second scan is taken after 10-15 minutes of  wash with PBS.
* Second scan is taken after 10-15 minutes of  wash with PBS.
[[File:EPFL2011_MITOMIchip_afterwashProt_illustration.png.png|300px| After PBS wash: GFP fluorescence ]]
[[File:EPFL2011_MITOMIchip_afterwashProt_illustration.png.png|300px| After PBS wash: GFP fluorescence ]]
[[File:EPFL2011_MITOMIchip_afterwashDNA_illustration.png|300px| After PBS wash: GFP fluorescence ]]
[[File:EPFL2011_MITOMIchip_afterwashDNA_illustration.png|300px| After PBS wash: GFP fluorescence ]]
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For data collection the fluorescence intensity measured under the button is normalised to the background and the DNA/protein ratio is calculated. This ratio is ploted against the intensity of the fluorescence of DNA in solution to make the saturation curves. And the Kd for each sequence is calculated from these curves.
 
[[File:EPFL2011_MITOMI_data_collection_points.JPG|400px]]
[[File:EPFL2011_MITOMI_data_collection_points.JPG|400px]]
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For data collection the fluorescence intensity measured under the button is normalised to the background and the DNA/protein ratio is calculated. This ratio is ploted against the intensity of the fluorescence of DNA in solution to make the saturation curves. And the Kd for each sequence is calculated from these curves by fitting the next equation:
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[[File:EPFL2011_MITOMI_Kd_fiting_formula.png|200px]]
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[[File:EPFL2011_MITOMI_Example_Saturation_curve_fited.png|400px]]
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{{:Team:EPF-Lausanne/Templates/Footer}}
{{:Team:EPF-Lausanne/Templates/Footer}}

Revision as of 23:40, 21 September 2011