Difference between revisions of "Part:BBa K4169030"

Revision as of 04:21, 12 October 2022

Cre-reverse loxP system:working when you need to express genes in the other direction

In the presence of theophylline, expression of subsequent Cre recombinase is induced.In addition, the promoter and RBS between the two loxP will be reversed, making it express genes in the other direction.

Usage and Biology

When our engineered bacteria feel theophylline, Cre expresses it, Cre recognizes the reverse repeat sequence at both ends of the loxP site and combines to form a dimer, which then binds to the dimer on another loxP site to form a tetramer. This way Cre mediates the reverse sequence reversal between loxPs.

Functional Parameters

To verify the functionality of our part, we had the upstream gene of the promoter replaced with red fluorescence，and then transferred our plasmid into Escherichia coli DH5α and measured the intensity of red fluorescence with a microplate reader. Unfortunately, our bacteria did not produce red fluorescence, and upon analysis, it was conjectured that the transcriptional loxP formed a strong secondary structure and could not translate the red fluorescence protein behind it.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 7
Illegal NheI site found at 30
Illegal NheI site found at 1292
Illegal NheI site found at 1315
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 454
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 39
Illegal AgeI site found at 140
1000
COMPATIBLE WITH RFC[1000]

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 <!-- Uncomment this to enable Functional Parameter display -->
 ===Functional Parameters===
-To verify the functionality of our part, We had the upstream gene of the promoter replaced with red fluorescence，and then transferred our plasmid into Escherichia coli DH5α and measured the intensity of red fluorescence with a microplate reader. Unfortunately, our bacteria did not produce red fluorescence, and after analysis, it may be that the transcriptional loxP forms a strong secondary structure and cannot translate the red fluorescent protein behind.
+To verify the functionality of our part, we had the upstream gene of the promoter replaced with red fluorescence，and then transferred our plasmid into Escherichia coli DH5α and measured the intensity of red fluorescence with a microplate reader. Unfortunately, our bacteria did not produce red fluorescence, and upon analysis, it was conjectured that the transcriptional loxP formed a strong secondary structure and could not translate the red fluorescence protein behind it.
 <partinfo>BBa_K4169030 parameters</partinfo>
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