Difference between revisions of "Part:BBa K2371012"
 
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In cell and molecular biology, the GFP gene is frequently used as a reporter of expression.It has been used in modified forms to make biosensors, and many animals have been created that express GFP, which demonstrates a proof of concept that a gene can be expressed throughout a given organism, in selected organs, or in cells of interest.  
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In cell and molecular biology, the GFP gene is often used as reporter.
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We add a T7 promotor and a RBS in front of the GFP in order to improve it to be utilized in our paired dCas9 T7 report system in cell free system together with N-t7-dCas9(BBa_K2371000) and C-t7-dCas9(BBa_K2371001).
  
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<h5>Figure1.PCR test of eforRed and amliGFP</h5>
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The user need to conduct a PCR to linearize the part before reacting with T7 report system.
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We conducted both PCR check(VF and VR) and enzyme check(XbaI and SpeI)
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[[File:BGIC-Union Part 123 GFP.png|700px]]
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<h5>Figure 1. The plasmid of pT7-amilGFP. pT7-amilGFP is composed of a T7 promotor, an RBS and the coding sequence of GFP. This plasmid is used to act as report gene in our report system.</h5>
  
 
[[File:Pcr test of eforRed and GFP.png|400px]]
 
[[File:Pcr test of eforRed and GFP.png|400px]]
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<h5>Figure 2. PCR check of eforRed and amliGFP. The expected result of eforRed should be around 1000bp. The insufficient specificity of primers produce another band but the band around 1000bp can present the success of construction. The expected result of amliGFP should be around 1000bp. The control is J23119 biobrick.</h5>
  
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<h5>Figure2.RD test of eforRed and amlGFP.</h5>
 
  
 
[[File:RD test of amliRed and GFP.png|400px]]
 
[[File:RD test of amliRed and GFP.png|400px]]
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<h5>Figure 3. RD check of eforRed and amliGFP. XbaI and SpeI were used. The expected length of both amliGFP and eforRed should be around 750bp.</h5>
 
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===Usage and Biology===
 
===Usage and Biology===

Latest revision as of 20:00, 1 November 2017


pT7-amilGFP


In cell and molecular biology, the GFP gene is often used as reporter. We add a T7 promotor and a RBS in front of the GFP in order to improve it to be utilized in our paired dCas9 T7 report system in cell free system together with N-t7-dCas9(BBa_K2371000) and C-t7-dCas9(BBa_K2371001).

The user need to conduct a PCR to linearize the part before reacting with T7 report system.

We conducted both PCR check(VF and VR) and enzyme check(XbaI and SpeI)

BGIC-Union Part 123 GFP.png

Figure 1. The plasmid of pT7-amilGFP. pT7-amilGFP is composed of a T7 promotor, an RBS and the coding sequence of GFP. This plasmid is used to act as report gene in our report system.

Pcr test of eforRed and GFP.png

Figure 2. PCR check of eforRed and amliGFP. The expected result of eforRed should be around 1000bp. The insufficient specificity of primers produce another band but the band around 1000bp can present the success of construction. The expected result of amliGFP should be around 1000bp. The control is J23119 biobrick.


RD test of amliRed and GFP.png

Figure 3. RD check of eforRed and amliGFP. XbaI and SpeI were used. The expected length of both amliGFP and eforRed should be around 750bp.

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 AgeI site found at 600
    Illegal AgeI site found at 712
  • 1000
    COMPATIBLE WITH RFC[1000]