Difference between revisions of "Part:BBa K1111013"
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<partinfo>BBa_K1111013 short</partinfo>
 
<partinfo>BBa_K1111013 short</partinfo>
  
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===Introduction===
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==Introduction==
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It is a fusion protein between an Ice Nucleation Protein (INP) and Streptavidin Alive made by Gibson Assembly.
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INP is an outer membrane protein found in the genome of Pseudomonas syringae, that is recognize by the E.coli protein secretion machinery.
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<!--The expression of this plasmid was tested by transforming cells and looking for microscopy at either of a fluorescent antibody again streptavidin or a fluorescent biotin (biotin and streptavidin have a strong affinity, their Kd is 10-15 M).-->
  
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INP sequence fused with Streptavidin Dead by gibson assembly.
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[[File:EPF-Lausanne map.jpg||thumb|400px|center|Figure 1: outline of the expected outcome of the plasmid]]
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The INP-streptavidin construct expresses streptavidin at the outer membrane of E.Coli. INP is an outer membrane protein foud in the genome of Pseudomonas syringae. It is under the control of Lac promoter and thus is constitutively expressed in E.Coli.
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The expression of this plasmid was tested by transforming cells and looking for microscopy at either of a fluorescent antibody again streptavidin or a fluorescent biotin (biotin and streptavidin have a strong affinity).
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==Usage and Biology==
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This part was designed to express streptavidin on the outer membrane of E.coli.
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<br> It serves as a negative control as the dead Streptavidin is though to not bind to biotin or with really decreased affinity.
  
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==Datasheet==
  
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===Usage and Biology===
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For further Characterization, refer to the datasheet below.
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We used this part as a negative control. We also have two other parts implying streptavidin non mutated, which are supposed to bien biotin (BBa_K1111012 and BBa_K1111014), and this streptavidin is mutated to decrease the Kd of the binding streptavidin-biotin. Thus, the streptavidin dead, although it is well exported to outer membrane, won't bind to biotin, or will bind weakly.
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[[File:Team-EPF-Lausanne_Sheet1.jpg|thumb|270px|left|CharacterizationSheet Page1 ]]
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[[File: Team-EPF-Lausanne_Sheet2.jpg|thumb|270px|left|CharacterizationSheet Page2 ]]
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<br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br>
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<br><br>[[File: Team-EPF-Lausanne_Sheet.pdf|thumb|270px|left|PDF link ]]
  
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<span class='h3bb'>Sequence and Features</span>
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==<span class='h3bb'>Sequence and Features</span>==
 
<partinfo>BBa_K1111013 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K1111013 SequenceAndFeatures</partinfo>
  

Latest revision as of 18:18, 4 October 2013

Ice Nucleation Protein fused to Streptavidin Dead

Introduction

It is a fusion protein between an Ice Nucleation Protein (INP) and Streptavidin Alive made by Gibson Assembly. INP is an outer membrane protein found in the genome of Pseudomonas syringae, that is recognize by the E.coli protein secretion machinery.

Figure 1: outline of the expected outcome of the plasmid

Usage and Biology

This part was designed to express streptavidin on the outer membrane of E.coli.
It serves as a negative control as the dead Streptavidin is though to not bind to biotin or with really decreased affinity.

Datasheet

For further Characterization, refer to the datasheet below.

CharacterizationSheet Page1
CharacterizationSheet Page2























File:Team-EPF-Lausanne Sheet.pdf

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal XbaI site found at 1727
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal XbaI site found at 1727
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal XbaI site found at 1727
    Illegal NgoMIV site found at 1036
    Illegal AgeI site found at 1691
    Illegal AgeI site found at 1742
  • 1000
    COMPATIBLE WITH RFC[1000]