Difference between revisions of "Part:BBa K629012"

 
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<partinfo>BBa_K629012 short</partinfo>
 
<partinfo>BBa_K629012 short</partinfo>
  
 
Used to test the chemotaxis when expression is controlled by a strong promoter.
 
Used to test the chemotaxis when expression is controlled by a strong promoter.
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== Background ==
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Polar localization is dependent on CheA-short. CheZ is a cytosolic phosphatase which functions in the chemotaxis signal transduction complex.
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Plays an important role in bacterial chemotaxis signal transduction pathway by accelerating the dephosphorylation of phosphorylated CheY (CheY-P).
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== Expression in ''E. Coli'' ==
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Before we started our functional test of CheZ, we have utilized western blot to determine whether there are enough soluble proteins in E. Coli. With 6×His tag on pET-32a, the result shows that CheZ has more expression with induction of IPTG.
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Note: Because the promoter is Plac here, the expression has a background, which means control also expresses some cheZ without induction of IPTG.
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[[Image:CheZ.jpg]]
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== Picture of Chemotaxis in ''cheZ'' knocked-out strain ==
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The main purpose of cheZ is to control chemotaxis in E. Coli. What we have done is to use different promoter to test how to get the better result from expression of CheZ. Below are pictures and parameters of chemotaxis test.
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Nalidixic acid Induction of promoter ''recAp''. If you need higher resolution pictures, please feel free to cantact us.
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[[Image:ddH2O.jpg]]
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[[Image:5 NAL.jpg]]
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[[Image:10 NAL.jpg]]
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<!-- Add more about the biology of this part here
 
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Revision as of 08:29, 9 October 2011

T7 promoter + lac operator + RBS +cheZ

Used to test the chemotaxis when expression is controlled by a strong promoter.


Background

Polar localization is dependent on CheA-short. CheZ is a cytosolic phosphatase which functions in the chemotaxis signal transduction complex. Plays an important role in bacterial chemotaxis signal transduction pathway by accelerating the dephosphorylation of phosphorylated CheY (CheY-P).


Expression in E. Coli

Before we started our functional test of CheZ, we have utilized western blot to determine whether there are enough soluble proteins in E. Coli. With 6×His tag on pET-32a, the result shows that CheZ has more expression with induction of IPTG.

Note: Because the promoter is Plac here, the expression has a background, which means control also expresses some cheZ without induction of IPTG.

CheZ.jpg


Picture of Chemotaxis in cheZ knocked-out strain

The main purpose of cheZ is to control chemotaxis in E. Coli. What we have done is to use different promoter to test how to get the better result from expression of CheZ. Below are pictures and parameters of chemotaxis test.

Nalidixic acid Induction of promoter recAp. If you need higher resolution pictures, please feel free to cantact us.

DdH2O.jpg

5 NAL.jpg

10 NAL.jpg


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal XbaI site found at 61
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal XbaI site found at 61
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal XbaI site found at 61
  • 1000
    COMPATIBLE WITH RFC[1000]