Difference between revisions of "Part:BBa K523025"

 
 
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__NOTOC__
 
__NOTOC__
 
<partinfo>BBa_K523025 short</partinfo>
 
<partinfo>BBa_K523025 short</partinfo>
  
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Fusion of exoglucanse (cex) and endoglucanase (cenA) from ''Cellulomonas fimi'' under the control of the Lac promoter.
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Lac promoter and RBS, followed by Cex exoglucanase from ''Cellulomonas fimi'' fused to CenA endoglucanase from the same species. The coding sequence contains, in order:
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* The GH10 exoglucanase domain
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* A (possibly inactivated) cellulose binding domain
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* The GH6 endoglucanase domain
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In the process of creating this part, we (Edinburgh 2011) tested putative correct clones on carboxymethylcellulose (CMC) plates, with Congo red dye. This test produces a zone of clearing if CMC is degraded by endoglucanase:
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[[Image:K523025_on_congo_red.jpg|400px|center]]
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Evidently we had three correct colonies, here numbered 8, 17, and 22. These had already been tested for their ability to degrade MUC, in an assay similar to that for <partinfo>BBa_K523016</partinfo>. Thus, both the exoglucanase and the endoglucanase domains have been shown to work.
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<span class='h3bb'>Sequence and Features</span>
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<span class='h3bb'>'''Sequence and Features'''</span>
 
<partinfo>BBa_K523025 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K523025 SequenceAndFeatures</partinfo>
  

Latest revision as of 18:52, 28 October 2011

Plac + RBS + cxnA (cex + cenA fusion)

Lac promoter and RBS, followed by Cex exoglucanase from Cellulomonas fimi fused to CenA endoglucanase from the same species. The coding sequence contains, in order:

  • The GH10 exoglucanase domain
  • A (possibly inactivated) cellulose binding domain
  • The GH6 endoglucanase domain


In the process of creating this part, we (Edinburgh 2011) tested putative correct clones on carboxymethylcellulose (CMC) plates, with Congo red dye. This test produces a zone of clearing if CMC is degraded by endoglucanase:


K523025 on congo red.jpg


Evidently we had three correct colonies, here numbered 8, 17, and 22. These had already been tested for their ability to degrade MUC, in an assay similar to that for BBa_K523016. Thus, both the exoglucanase and the endoglucanase domains have been shown to work.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NotI site found at 1148
    Illegal NotI site found at 2846
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 2759
    Illegal XhoI site found at 2257
    Illegal XhoI site found at 2506
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 781
    Illegal NgoMIV site found at 1154
    Illegal NgoMIV site found at 1656
    Illegal NgoMIV site found at 2037
    Illegal NgoMIV site found at 2962
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 1201
    Illegal SapI.rc site found at 1284