Difference between revisions of "Part:BBa K4165167"
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This composite part consists of T7 promoter (BBa_K3633015), lac operator (BBa_K4165062), RBS 4 (BBa_K4165264), 6x His-tag (BBa_K4165020), DocS (BBa_K3396000) and T7 terminator (BBa_K731721).
 
This composite part consists of T7 promoter (BBa_K3633015), lac operator (BBa_K4165062), RBS 4 (BBa_K4165264), 6x His-tag (BBa_K4165020), DocS (BBa_K3396000) and T7 terminator (BBa_K731721).
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===Source===
 
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Synthesized
 
  
 
===Usage and Biology===
 
===Usage and Biology===
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<partinfo>BBa_K4165167 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K4165167 SequenceAndFeatures</partinfo>
  
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===Dry Lab===
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===Modeling===
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<p style=" font-weight: bold; font-size:14px;"> Modeling </p>
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Model “peptide (His_DocS) modeled by TRrosetta model1” Ranked the Third model from our 4 ones with a score 5 out of 6 and values of  
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Model “peptide (His_DocS) modeled by TRrosetta model1” Ranked Third model from our 4 ones with score 5 out of 6 and values of C Beta deviasion = 0 , clash score = 0 , molprobitey = 0.77 , Ramachandran favored = 96 , Ramachandran outlier = 0 , Q Mean_4 = -0.29 , Q Mean_6 = -0.27
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<html>
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<style>
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table, th, td {
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  border:1px solid black; margin-left:auto;margin-right:auto;
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}
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</style>
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<body>
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<table style="width:65%">
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<table>
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  <tr>
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    <th>cbeta_deviations</th>
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    <th>clashscore</th>
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    <th>molprobity</th>
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    <th>ramachandran_favored</th>
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    <th>ramachandran_outliers</th>
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    <th>Qmean_4</th>
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    <th>Qmean_6</th>
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  </tr>
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  <tr>
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    <td>0</td>
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    <td>0</td>
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    <td>0.77</td>
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    <td>96</td>
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    <td>0</td>
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    <td>-0.29</td>
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    <td>-0.27</td>
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  </tr>
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</table>
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</body>
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</html>
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<html>
 
<html>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/dry-lab/mathematical-modeling/mathematical-modeling/his-doc.png" style="margin-left:200px;" alt="" width="500" /></p>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/parts-registry/his-docs.jpg" style="margin-left:200px;" alt="" width="500" /></p>
 
</html>
 
</html>
  
  
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                          Figure 1. The 3D structure of His Docs model visualized by Pymol.
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                              Figure 1. The 3D structure of His Docs model Displayed on Pymol.
  
 
<p style=" font-weight: bold; font-size:14px;"> Mathematical modeling </p>
 
<p style=" font-weight: bold; font-size:14px;"> Mathematical modeling </p>
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<html>
 
<html>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/dry-lab/mathematical-modeling/mathematical-modeling/his-doc2.png" alt="" width="500" /></p>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/dry-lab/mathematical-modeling/his-doc2.png"style="margin-left:200px;" alt="" width="500" /></p>
 
</html>
 
</html>
  
  
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                  Figure 2. this figure shows the results from the transcription and translation code showing  
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              Figure 2. this figure shows the results from the transcription and translation code showing  
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                  the variation of mRNA and protein concentrations with time compared with the wet lab results.
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              the variation of mRNA and protein concentrations with time compared with the wet lab results.
 
   
 
   
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===WetLab Results===
 
===WetLab Results===
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<p style=" font-weight: bold; font-size:14px;"> Transformation of His Doc in BL-21 using pGS-21a vector </p>
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<p style=" font-weight: bold; font-size:14px;"> Comparison between the mathematical model of dry lab and the wet lab results of His DOC using sonicator and chemical lysis </p>
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We have made a statistical analysis (t-test) to investigate the mathematical model's effectiveness in predicting the amount of the produced protein due to IPTG induction, whereas the test compares the mathematical model expected results to the replicates made in the induction experiment, given that all the condition of the experiment has been input to the model generating the predicted results.
 
<html>
 
<html>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/parts-registry/wetlab-results/his-doc-pgs.jpg" style="margin-left:200px;" alt="" width="500" /></p>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/data-analysis/tir-wet-expression/his-doc-son.jpg" style="margin-left:200px;" alt="" width="500" /></p>
 
</html>
 
</html>
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                                    Figure 3. Transformed plate of His Doc + pGS-21a
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              Figure 3. This graph shows the correlation between the mathematical model of the dry lab and the wet lab
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<p style=" font-weight: bold; font-size:14px;"> Transformation of His Doc in DH-5 alpha using pJET vector </p>
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                                              results of the His DOC using a sonicator
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<html>
 
<html>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/parts-registry/wetlab-results/his-doc-pjet.jpg" style="margin-left:200px;" alt="" width="500" /></p>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/data-analysis/tir-wet-expression/his-docs-chem.jpg" style="margin-left:200px;" alt="" width="500" /></p>
 
</html>
 
</html>
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                                    Figure 4. Transformed plate of His Doc + pJET
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              Figure 4. This graph shows the correlation between the mathematical model of the dry lab and the wet lab
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<p style=" font-weight: bold; font-size:14px;"> Comparison between chemical lysis and sonication for His DOC </p>
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                                                results of His DOC using chemical lysis
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<html>
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<p><img src="https://static.igem.wiki/teams/4165/wiki/data-analysis/sonication-or-chemical/sonication-or-chemical/his-docs.jpg" style="margin-left:200px;" alt="" width="500" /></p>
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</html>
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Figure 5. This graph shows the difference between chemical lysis and sonication for His DOC, after we had the results we optimized our protocol to use chemical lysis for His DOC
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<!-- Uncomment this to enable Functional Parameter display  
 
<!-- Uncomment this to enable Functional Parameter display  

Latest revision as of 23:50, 13 October 2022


His - DocS

This composite part consists of T7 promoter (BBa_K3633015), lac operator (BBa_K4165062), RBS 4 (BBa_K4165264), 6x His-tag (BBa_K4165020), DocS (BBa_K3396000) and T7 terminator (BBa_K731721).

Usage and Biology

DOCs protein is a part of the Snitch system, which binds to the His tag for the purification of the Docs.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 45

Modeling

Model “peptide (His_DocS) modeled by TRrosetta model1” Ranked the Third model from our 4 ones with a score 5 out of 6 and values of

cbeta_deviations clashscore molprobity ramachandran_favored ramachandran_outliers Qmean_4 Qmean_6
0 0 0.77 96 0 -0.29 -0.27 



                             Figure 1. The 3D structure of His Docs model Displayed on Pymol.

Mathematical modeling

Transcription rate and translation rate under T7 promotor

the mathematical modeling was based on our code for the calculation of transcription and translation (you can find it in the code section) beside with the estimated results from the wet lab. 



              Figure 2. this figure shows the results from the transcription and translation code showing 
              the variation of mRNA and protein concentrations with time compared with the wet lab results.

WetLab Results

Comparison between the mathematical model of dry lab and the wet lab results of His DOC using sonicator and chemical lysis

We have made a statistical analysis (t-test) to investigate the mathematical model's effectiveness in predicting the amount of the produced protein due to IPTG induction, whereas the test compares the mathematical model expected results to the replicates made in the induction experiment, given that all the condition of the experiment has been input to the model generating the predicted results. 

              Figure 3. This graph shows the correlation between the mathematical model of the dry lab and the wet lab 
                                              results of the His DOC using a sonicator


              Figure 4. This graph shows the correlation between the mathematical model of the dry lab and the wet lab 
                                               results of His DOC using chemical lysis