Difference between revisions of "Part:BBa K2407014"

 
 
Line 3: Line 3:
 
<partinfo>BBa_K2407014 short</partinfo>
 
<partinfo>BBa_K2407014 short</partinfo>
  
−
loading
+
<h3>Design</h3>
 +
<hr>
  
 +
<p>This part was improved from <partinfo>BBa_K2407000</partinfo> and the source can be tracked back to <partinfo>BBa_K2165004</partinfo>.</p>
 +
 +
<p>In our experiment, we noticed that <i>CUP1</i> promoter still has a certain degree of leakage expression. To make a better biosensor, we planned to reduce the leakage expression and increase the sensitivity. To reach this goal, we took the fluorescence intensity at both induction or not into evaluation indexes.</p>
 +
 +
<p>The technology of error-prone PCR was taken into our experiment. Although there are many methods to introduce genetic diversity into a parent sequence, error-prone PCR is the most common way of creating a combinatorial library based on a single sequence. By adding some heavy metal ions into the PCR buffer and preparing dNTPs with different composition, new mutants were introduced into <i>CUP1</i> promoter.</p>
 +
 +
<p>This is one of the mutants from <i>CUP1</i> promoter, which was named with EP-5. The changes in sequence were uploaded in the "sequence and feature" part.</p>
 +
 +
<h3>Characterization</h3>
 +
<hr>
 +
<p>We tested the leakage expression and response rages of each selected mutants, and results were shown below.</p>
 +
 +
[[File:Tianjin-Demonstrate-4-5-leakage.png|700px|thumb|center|'''Fig 1'''.The leakage expression of different promoters ]]
 +
 +
[[File:Tianjin-Demonstrate-4-6-EP.png|700px|thumb|center|'''Fig 2'''.Different biosensors showed different sensitivities and response rages ]]
 +
 +
<p>Visit <partinfo>BBa_K2407013</partinfo>, <partinfo>BBa_K2407015</partinfo>, and <partinfo>BBa_K2407016</partinfo> to see other mutants.</p>
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here
 
===Usage and Biology===
 
===Usage and Biology===

Latest revision as of 12:10, 27 October 2017


a modified CUP1 promoter obtained by Error-Prone PCR EP5

Design

This part was improved from BBa_K2407000 and the source can be tracked back to BBa_K2165004.

In our experiment, we noticed that CUP1 promoter still has a certain degree of leakage expression. To make a better biosensor, we planned to reduce the leakage expression and increase the sensitivity. To reach this goal, we took the fluorescence intensity at both induction or not into evaluation indexes.

The technology of error-prone PCR was taken into our experiment. Although there are many methods to introduce genetic diversity into a parent sequence, error-prone PCR is the most common way of creating a combinatorial library based on a single sequence. By adding some heavy metal ions into the PCR buffer and preparing dNTPs with different composition, new mutants were introduced into CUP1 promoter.

This is one of the mutants from CUP1 promoter, which was named with EP-5. The changes in sequence were uploaded in the "sequence and feature" part.

Characterization

We tested the leakage expression and response rages of each selected mutants, and results were shown below.

Fig 1.The leakage expression of different promoters
Fig 2.Different biosensors showed different sensitivities and response rages

Visit BBa_K2407013, BBa_K2407015, and BBa_K2407016 to see other mutants.

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
    COMPATIBLE WITH RFC[1000]