Difference between revisions of "Part:BBa K2922032"
 
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The usage of T7 promoter and RBS is to highly express the Colicin-N immunity protein.
 
The usage of T7 promoter and RBS is to highly express the Colicin-N immunity protein.
 
===Identification===
 
===Identification===
−
When we are building this circuit, we are doing the nucleic acid gel electrophoresis experiment to verify. After the circuit is built, we send the plasmid to sequence, and get the correct sequencing.
+
When we were building this circuit, we did the nucleic acid gel electrophoresis experiment to verify. After the circuit was built, we sent the plasmid to sequence, and got the correct sequencing.
−
After new molecular cloning experiments, we do Enzyme-Cut identification to certify the plasmid is correct. We use the <i>Eco</i>RI and <i>Pst</i>I to cut the plasmid, then we get the target separate fragment-542bp. (Fig.1)
+
After new molecular cloning experiments, we did Enzyme-Cut identification to certify the plasmid was correct. We used the <i>Eco</i>RI and <i>Pst</i>I to cut the plasmid, then we got the target separate fragment-462bp. (Fig.1)
 
<table><tr><th>[[File:T7+RBS+Nimm.png|thumb|300px|Fig.1 The result of this plasmid cut with enzyme <i>Eco</i>RI and <i>Pst</i>I.]]</th><th></table>
 
<table><tr><th>[[File:T7+RBS+Nimm.png|thumb|300px|Fig.1 The result of this plasmid cut with enzyme <i>Eco</i>RI and <i>Pst</i>I.]]</th><th></table>
−
The plasmids are transformed into BL21(DE3).After growing the single colonies,SDS-PAGE protein gel electrophoresis is used to certify the expression of Ekil,then we get the target molecular weight-15kDa.(Fig.2)
+
The plasmid was transformed into BL21(DE3).After growing the single colonies,SDS-PAGE protein gel electrophoresis was used to certify the expression of Nimm,then we got the target molecular weight-15kDa.(Fig.2)
−
<table><tr><th>[[File:Nimm protein.png|thumb|300px|Fig.1 The result of the protein molecular weight.]]</th><th></table>
+
<table><tr><th>[[File:Nimm protein.png|thumb|720px|Fig.1 The result of the protein molecular weight.]]</th><th></table>
  
 
===Sequence and Features===
 
===Sequence and Features===

Latest revision as of 18:06, 20 October 2019


T7 promoter-RBS(B0034)_ cni(Nimm)

Summary

The usage of T7 promoter and RBS is to highly express the Colicin-N immunity protein.

Identification

When we were building this circuit, we did the nucleic acid gel electrophoresis experiment to verify. After the circuit was built, we sent the plasmid to sequence, and got the correct sequencing. After new molecular cloning experiments, we did Enzyme-Cut identification to certify the plasmid was correct. We used the EcoRI and PstI to cut the plasmid, then we got the target separate fragment-462bp. (Fig.1)

Fig.1 The result of this plasmid cut with enzyme EcoRI and PstI.

The plasmid was transformed into BL21(DE3).After growing the single colonies,SDS-PAGE protein gel electrophoresis was used to certify the expression of Nimm,then we got the target molecular weight-15kDa.(Fig.2)

Fig.1 The result of the protein molecular weight.

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]