Difference between revisions of "Part:BBa K3396016"
 
(3 intermediate revisions by 3 users not shown)
Line 1: Line 1:
  
 
__NOTOC__
 
__NOTOC__
−
<partinfo>BBa_K3396008 short</partinfo>
+
<partinfo>BBa_K3396016 short</partinfo>
  
−
A dual luciferase reporter to monitor the Fluc tagged protein abundance while rluc can be used to normalize irrelevant factors.
+
Dual luciferase reporter plasmid to indicate the target protein abundance. The coding sequence of target protein can be inserted into the replaceable region with Golden Gate assembly, the ratio Fluc/Rluc shall indicate the normalized protein amount of the target.
  
 
===Usage and Biology===
 
===Usage and Biology===
Line 9: Line 9:
  
 
===Special Design===
 
===Special Design===
−
Special designs were taken to optimize the applicability and adaptivity of such parts. Specifically, a novel designed substitution system, through which, target proteins could be fused with Fluc fragment using Golden-Gate Assembly, was introduced to dramatically simplify the cloning process.
+
Special designs were taken to simplify the uses of this part. Specifically, a novel designed substitution system, through which, target proteins could be easily fused with Fluc fragment using Golden-Gate Assembly, was introduced to dramatically simplify the cloning process.
  
 +
===Characterization===
 +
We use Golden-Gate Assembly to fuse GFP with Fluc fragment. This system can be used to quantify the abundance of specific target protein. Then, we compared the degradation curves of GFP and Fluc as reporter molecules, respectively and result showed that the detection of Fluc is more accurate.
 +
 +
===Result===
 +
We use Golden-Gate Assembly to fuse GFP with Fluc fragment. This system can be used to quantify the abundance of specific target protein. Then, we compared the degradation curves of GFP and Fluc as reporter molecules, respectively and result showed that the detection of Fluc is more accurate.
 +
 +
<html>
 +
<p><img src="https://static.igem.org/mediawiki/parts/3/3b/T--NUDT_CHINA--BBa_K3396016_Fig1.png" alt="" width="500" /></p>
 +
</html>
 +
 +
 +
Figure 1.Detection accuracy of different reporter system.
 +
 +
 +
===Sequence and Features===
  
 
<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
−
<partinfo>BBa_K3396008 SequenceAndFeatures</partinfo>
+
<partinfo>BBa_K3396016 SequenceAndFeatures</partinfo>
  
  
 
<!-- Uncomment this to enable Functional Parameter display  
 
<!-- Uncomment this to enable Functional Parameter display  
 
===Functional Parameters===
 
===Functional Parameters===
−
<partinfo>BBa_K3396008 parameters</partinfo>
+
<partinfo>BBa_K3396016 parameters</partinfo>
 
<!-- -->
 
<!-- -->

Latest revision as of 18:34, 27 October 2020


CMV-replaceable-Fluc-P2A-Rluc

Dual luciferase reporter plasmid to indicate the target protein abundance. The coding sequence of target protein can be inserted into the replaceable region with Golden Gate assembly, the ratio Fluc/Rluc shall indicate the normalized protein amount of the target.

Usage and Biology

BBa_K3396008 is a Dual Luciferase Reporter system to quantify the abundance of specific target protein. Dual luciferase assay was introduced to normalize the differences caused by irrelevant factors.

Special Design

Special designs were taken to simplify the uses of this part. Specifically, a novel designed substitution system, through which, target proteins could be easily fused with Fluc fragment using Golden-Gate Assembly, was introduced to dramatically simplify the cloning process.

Characterization

We use Golden-Gate Assembly to fuse GFP with Fluc fragment. This system can be used to quantify the abundance of specific target protein. Then, we compared the degradation curves of GFP and Fluc as reporter molecules, respectively and result showed that the detection of Fluc is more accurate.

Result

We use Golden-Gate Assembly to fuse GFP with Fluc fragment. This system can be used to quantify the abundance of specific target protein. Then, we compared the degradation curves of GFP and Fluc as reporter molecules, respectively and result showed that the detection of Fluc is more accurate.


Figure 1.Detection accuracy of different reporter system.


Sequence and Features

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
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 871
    Illegal NgoMIV site found at 2215
    Illegal NgoMIV site found at 2236
    Illegal AgeI site found at 1939
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 3102
    Illegal SapI.rc site found at 2121