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<p style="font-family: quicksand;font-size:150%;">RNA thermodynamic Modelling</p> | <p style="font-family: quicksand;font-size:150%;">RNA thermodynamic Modelling</p> | ||
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+ | <center><img src="https://static.igem.org/mediawiki/2017/9/90/Toehold_Switch_Model.jpeg" width="100%" height="auto" class=" igem-logo"></center> | ||
+ | <p style="font-family: roboto;font-size:115%;"> | ||
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<p style="font-family: roboto;font-size:125%;color:blue"><b>Assumption 1: Switch MFE (Minimum Free Energy) correlates with the expression leakage</p></b> | <p style="font-family: roboto;font-size:125%;color:blue"><b>Assumption 1: Switch MFE (Minimum Free Energy) correlates with the expression leakage</p></b> | ||
<p style="font-family: roboto;font-size:115%;"> | <p style="font-family: roboto;font-size:115%;"> | ||
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<p style="font-family: quicksand;font-size:150%;">Suboptimal Structure Modelling</p> | <p style="font-family: quicksand;font-size:150%;">Suboptimal Structure Modelling</p> | ||
− | <p style="font-family: roboto;font-size:125%;color | + | <p style="font-family: roboto;font-size:125%;color:blue"><b>Assumption 4: Accessibility of toehold domain correlates with the performance of switch</p></u> |
<p style="font-family: roboto;font-size:115%;"> | <p style="font-family: roboto;font-size:115%;"> | ||
● The toehold domain (first 15 nts) of the switch RNA is crucial for the binding of the trigger RNA to the switch RNA to initiate the switch-unwinding process: | ● The toehold domain (first 15 nts) of the switch RNA is crucial for the binding of the trigger RNA to the switch RNA to initiate the switch-unwinding process: | ||
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Toehold domain must have minimal paired bases in the switch RNA to ensure the successful binding of this domain with the complementary sequence in the trigger RNA, which allows the unwinding of the switch RNA and permits translation of the reporter protein, RFP, to occur. </p> | Toehold domain must have minimal paired bases in the switch RNA to ensure the successful binding of this domain with the complementary sequence in the trigger RNA, which allows the unwinding of the switch RNA and permits translation of the reporter protein, RFP, to occur. </p> | ||
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<p style="font-family: roboto;font-size:115%;"> | <p style="font-family: roboto;font-size:115%;"> | ||
− | From the table above, we observed that only 3 out of 15 of our predictions were incorrect, showing promising accuracy of this prediction method. None of the 3 incorrect predictions were false negatives, indicating that the suboptimal structure prediction method should be mostly applied to filtering out switch sequences with low predicted performances.</p> | + | From the table above, we observed that only <em>3 out of 15 of</em> our predictions were incorrect, showing promising accuracy of this prediction method. None of the 3 incorrect predictions were false negatives, indicating that <strong>the suboptimal structure prediction method should be mostly applied to filtering out switch sequences with low predicted performances</strong>.</p> |
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− | <p style="font-family: quicksand;font-size:150%;"> | + | <p style="font-family: quicksand;font-size:150%;">Experimental verification of the thermodynamic mode</p> |
<p style="font-family: roboto;font-size:125%;"><u><b> Correlation of switch MFE (Minimum Free Energy) and expression leakage </p></b></u> | <p style="font-family: roboto;font-size:125%;"><u><b> Correlation of switch MFE (Minimum Free Energy) and expression leakage </p></b></u> | ||
<img src="https://static.igem.org/mediawiki/2017/1/1e/CUHK_SMFE.jpg" width="40%" height="auto" class=" igem-logo"> | <img src="https://static.igem.org/mediawiki/2017/1/1e/CUHK_SMFE.jpg" width="40%" height="auto" class=" igem-logo"> | ||
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<img src="https://static.igem.org/mediawiki/2017/c/c1/CUHK_MFED.jpg" width="40%" height="auto" class=" igem-logo"> | <img src="https://static.igem.org/mediawiki/2017/c/c1/CUHK_MFED.jpg" width="40%" height="auto" class=" igem-logo"> | ||
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− | We initially thought that the larger the MFE difference between the sum of the MFE of switch and trigger and that of the switch-trigger duplex, the more favourable the duplex structure will be. However, there seems to be a weak positive correlation between the expression level of the switch-trigger duplex and the MFE difference of a switch. This could be because that a low MFE difference could indicate a more similar conformation of the switch/trigger RNA to the switch-trigger duplex RNA. This could lead to less conformational changes in the duplex formation process, making the process faster. | + | We initially thought that the larger the MFE difference between the sum of the MFE of switch and trigger and that of the switch-trigger duplex, the more favourable the duplex structure will be. However, there seems to be a weak positive correlation between the expression level of the switch-trigger duplex and the MFE difference of a switch. This could be because that a low MFE difference could indicate a more similar conformation of the switch/trigger RNA to the switch-trigger duplex RNA. This could lead to less conformational changes in the duplex formation process, making the process faster. Thus, a small MFE difference could lead to an increased expression level of the switch-trigger duplex. |
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<img src="https://static.igem.org/mediawiki/2017/3/33/CUHK_bp.jpg" width="40%" height="auto" class=" igem-logo"> | <img src="https://static.igem.org/mediawiki/2017/3/33/CUHK_bp.jpg" width="40%" height="auto" class=" igem-logo"> | ||
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− | The plot shows some kind of negative correlation between RFP signal in the presence of trigger and the number of base pairs in the toehold domain. Some exceptions were observed, they are N1-3, N9-3, N9-1. H5-3 and N1-2. This supports the | + | The plot shows some kind of negative correlation between RFP signal in the presence of trigger and the number of base pairs in the toehold domain. Some exceptions were observed, they are N1-3, N9-3, N9-1. H5-3 and N1-2. This supports the assumption in the suboptimal structure modelling that an increasing number of unpaired bases in the toehold domain promotes initial binding of a trigger RNA to the corresponding switch RNA. |
Latest revision as of 04:01, 2 November 2017