Difference between revisions of "Part:BBa K2922031"

Latest revision as of 18:04, 20 October 2019

T7 promoter-RBS(B0034)_kil(Ekil)

Summary

The usage of T7 promoter and RBS is to highly express the Lysis protein for Colicin-E1.

Identification

When we wre 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-204bp. (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 Ekil,then we got the target molecular weight-4.8kDa.(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
INCOMPATIBLE WITH RFC[21]
Unknown
23
INCOMPATIBLE WITH RFC[23]
Unknown
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 176
1000
COMPATIBLE WITH RFC[1000]

@@ Line 7: / Line 7: @@
 ===Identification===
 When we wre 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 <i>Eco</i>RI and <i>Pst</i>I to cut the plasmid, then we got the target separate fragment-284bp. (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-204bp. (Fig.1)
 <table><tr><th>[[File:TEimm TEkil.png|thumb|720px|Fig.1 The result of this plasmid cut with enzyme <i>Eco</i>RI and <i>Pst</i>I.]]</th><th></table>
 The plasmid was transformed into BL21(DE3).After growing the single colonies,SDS-PAGE protein gel electrophoresis was used to certify the expression of Ekil,then we got the target molecular weight-4.8kDa.(Fig.2)