Difference between revisions of "Part:BBa K1316007"
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<h3>Characterisation</h3>
 
<h3>Characterisation</h3>
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<p>
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Different type of experiments were conducted to characterize the Landmine Detection BioBricks.
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</p>
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<br>
  
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Adding these genes did unfortunately show no clear improvement on the response of the ''yqjF'' promoter in front of 2,4-DNT (Data not shown).
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<p>
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The different constructs used are:
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<ul>
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<li> p[F]::mKATE, also referred here as LD2  </li>
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<li> p[J]::mKATE, also referred here as LD3  </li>
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<li> p[F] incl. N-enzymes, also referred here  as LD4  </li>
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<li> p[J] incl. N-enzymes, also referred here as LD5  </li>
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<li> p[F]::mKATE p[J]::mKATE, also referred here  as LD6  </li>
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</ul>
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</p>
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<br>
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<h3> Plate Reader </h3>
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<p>
  
  

Revision as of 11:53, 16 October 2014

yqjF promoter coupled to mKate2 reporter gene and Rhamnose promoter coupled to N-genes

Promoter of the yqjF gene, which is activated by the presence of several nitrogen-based compounds such as 2,4,6-TNT, 2,4-DNT and 1,3-DNB. mKate2 is a far-red fluorescent protein used with reporting purposes. mKate2 codon usage is optimised for high expression in mammalian cells (humanised). It is, nevertheless, suitable for propagation in E. coli.

nfsA, nfsB and nemA (refered here as N-genes) are the genes for NfsA, NfsB and NEM reductases, which presumably play a role in 2,4-DNT and 2,4,6-TNT metabolism. These genes are regulated by the inducible Rhamnose promoter This construct is, then, designed to be able to detect and quantify the presence of compunds such as 2,4,6-TNT, 2,4-DNT and 1,3-DNB. The presence of the N-genes aims to enhance this response.


Characterisation

Different type of experiments were conducted to characterize the Landmine Detection BioBricks.


The different constructs used are:

  • p[F]::mKATE, also referred here as LD2
  • p[J]::mKATE, also referred here as LD3
  • p[F] incl. N-enzymes, also referred here as LD4
  • p[J] incl. N-enzymes, also referred here as LD5
  • p[F]::mKATE p[J]::mKATE, also referred here as LD6


Plate Reader

Sequence and Features

Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 128
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 733
    Illegal BsaI.rc site found at 922