Difference between revisions of "Part:BBa K2324012"
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This part contains the <i> fim operon </i>, minus the <i>fimH</i> under control of a constitutive promoter. The constituent genes code for structural proteins, such as FimA which makes up the majority of the main body of a pilus, and others which support the process of pilus biogenesis. When co-transformed with FimH constructs from our project, which have rhamnose-inducible promoters, modified type I pili may be produced. The part requires no induction, and therefore is able to continuously produce the large mass of protein encoded in the operon.  
 
This part contains the <i> fim operon </i>, minus the <i>fimH</i> under control of a constitutive promoter. The constituent genes code for structural proteins, such as FimA which makes up the majority of the main body of a pilus, and others which support the process of pilus biogenesis. When co-transformed with FimH constructs from our project, which have rhamnose-inducible promoters, modified type I pili may be produced. The part requires no induction, and therefore is able to continuously produce the large mass of protein encoded in the operon.  
  
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<!-- Add more about the biology of this part here
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<h2>Results</h2>
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===Usage and Biology===
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We transformed this part into <i>E. coli</i> Top10 to image them using a negative staining method on a TEM to obtain an electron micrograph to visualise pili.
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<html><body>
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<img src="https://static.igem.org/mediawiki/2017/a/af/T--Exeter--Top10J23NS.jpeg" height="475px" width="750px"/>
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</body></html>
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<b>Figure 1</b>
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Top10 with the <i>fim operon</i> under the control of promoter P_J23100 showed no visible signs of pili expression with insertion of the operon alone.
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<html><body>
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<img src="https://static.igem.org/mediawiki/2017/4/41/T--Exeter--FimBKOj23.jpeg" height="475px" width="750px"/>
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</body></html>
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<b>Figure 2</b>
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We transformed a plasmid containing the <i>fim operon</i> under control of promoters P_J23100 into &#916;FimB. It showed flagella on their cell surface, but no pili on the surface of the cell.
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<h2>Conclusion</h2>
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We were unsuccessful in producing pili on the cell surface of the cell, however the operon was not co-transformed with a <i>fimH</i> plasmid. 
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K2324012 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K2324012 SequenceAndFeatures</partinfo>

Revision as of 22:25, 1 November 2017


pJ23100_FimOp

This part contains the fim operon , minus the fimH under control of a constitutive promoter. The constituent genes code for structural proteins, such as FimA which makes up the majority of the main body of a pilus, and others which support the process of pilus biogenesis. When co-transformed with FimH constructs from our project, which have rhamnose-inducible promoters, modified type I pili may be produced. The part requires no induction, and therefore is able to continuously produce the large mass of protein encoded in the operon.

Results

We transformed this part into E. coli Top10 to image them using a negative staining method on a TEM to obtain an electron micrograph to visualise pili.

Figure 1 Top10 with the fim operon under the control of promoter P_J23100 showed no visible signs of pili expression with insertion of the operon alone.

Figure 2 We transformed a plasmid containing the fim operon under control of promoters P_J23100 into ΔFimB. It showed flagella on their cell surface, but no pili on the surface of the cell.

Conclusion

We were unsuccessful in producing pili on the cell surface of the cell, however the operon was not co-transformed with a fimH plasmid.



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
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 2648
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 931
    Illegal AgeI site found at 962
  • 1000
    COMPATIBLE WITH RFC[1000]