Difference between revisions of "Part:BBa K1202101:Experience"

 
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We need to measure enzyme activity with observing the AI-2 production from SAH by enzyme .
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It’s possible to measure the autoinducer concentration by ellman’s assay.
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Ellmans assay needs liquid culture of enzyme expressing bacteria so we preapare liquid cultures (with antibiotics) of them. After incubating liquid cultures add 0,1 mM substrate (SAH in 10X PBS containing %1 BSA) and incubated SAH added liquid cultures at room temprature for two hour. While the bacteria incubating, enzymes producted autoinducer 2. After incubating, santrifujed and measure the supernatants. We measure autoinducer concentration, enzymes production. We use transformed bacteria as a negative control.
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Ellman’s assay result:
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[[File:Aı.jpg]]
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As you see in the grafic, His-Enzyme complex and ezyme complex produced Auto Inducer-2 from the SAH.
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This result telling us enzymes are expressed by E.coli and succesfully produced autoinducer-2.
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For more information, go to Result/Discussion page
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To observe inducible promoters response, we co incubated two different bacteria. The first bacteria culture is expressing enzyme system and the second culture is including inducible promoter.
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[[File:DSC 0257.jpg]]
  
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__NOTOC__
 
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This experience page is provided so that any user may enter their experience using this part.<BR>Please enter
 
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how you used this part and how it worked out.
 
  
 
===Applications of BBa_K1202101===
 
===Applications of BBa_K1202101===

Revision as of 03:28, 5 October 2013

We need to measure enzyme activity with observing the AI-2 production from SAH by enzyme . It’s possible to measure the autoinducer concentration by ellman’s assay. Ellmans assay needs liquid culture of enzyme expressing bacteria so we preapare liquid cultures (with antibiotics) of them. After incubating liquid cultures add 0,1 mM substrate (SAH in 10X PBS containing %1 BSA) and incubated SAH added liquid cultures at room temprature for two hour. While the bacteria incubating, enzymes producted autoinducer 2. After incubating, santrifujed and measure the supernatants. We measure autoinducer concentration, enzymes production. We use transformed bacteria as a negative control. Ellman’s assay result: Aı.jpg As you see in the grafic, His-Enzyme complex and ezyme complex produced Auto Inducer-2 from the SAH. This result telling us enzymes are expressed by E.coli and succesfully produced autoinducer-2.

For more information, go to Result/Discussion page

To observe inducible promoters response, we co incubated two different bacteria. The first bacteria culture is expressing enzyme system and the second culture is including inducible promoter. File:DSC 0257.jpg


Applications of BBa_K1202101

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