Team:EPF-Lausanne/Our Project/TetR mutants/MITOMI data

From 2011.igem.org

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(In Vitro Characterization)
(E37A W43S T141A)
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===In Vitro Characterization===
===In Vitro Characterization===
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Using the <html> <a href="https://2011.igem.org/Team:EPF-Lausanne/Tools/MITOMI">MITOMI</a></html> technique we determined the DNA binding landscape of the TetR E37A  W43S T141A  mutant. To do so, first we designed and generated the library of double stranded DNA sequences that cover all possible single base substitution within the tetO operator sequence. Based on that library we measured the dissociation constants of the mutant to variable tetO-like sequences and determined the specificity of the mutant to the tet operator sequence (expressed as a PWM). We compared the measured DNA binding affinities of the E37A W43S T141A mutant to the data for the wt-TetR and found that E37A W43S T141A mutant has a lower affinity to the tetO.  
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Using the <html> <a href="https://2011.igem.org/Team:EPF-Lausanne/Tools/MITOMI">MITOMI</a></html> technique we determined the DNA binding landscape of the TetR E37A  W43S T141A  mutant. To do so, first we designed and generated the library of double stranded DNA sequences that cover all possible single base substitution within the tetO operator sequence. Based on that library we measured the dissociation constants of the mutant to variable tetO-like sequences and determined the specificity of the mutant to the tet operator sequence (expressed as a PWM). The measured DNA binding affinities of the E37A W43S T141A mutant show that these mutations alter the sequence recognition, while the symmetry and the spacer affinities are conserved.

Revision as of 03:23, 22 September 2011