Difference between revisions of "Part:BBa K1202110:Experience"

 
 
(4 intermediate revisions by 2 users not shown)
Line 1: Line 1:
 +
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.
  
−
__NOTOC__
+
Ellman’s assay result:
−
This experience page is provided so that any user may enter their experience using this part.<BR>Please enter
+
 
−
how you used this part and how it worked out.
+
[[File: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.
 +
 
 +
 
 +
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:Co-incubation.png]]
 +
 
 +
We saw the red color at the pellet. So, it means the enzyme system is working and inducible promoter is succesfully induced from AI-2.
  
 
===Applications of BBa_K1202110===
 
===Applications of BBa_K1202110===

Latest revision as of 10:43, 9 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.


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.

Co-incubation.png

We saw the red color at the pellet. So, it means the enzyme system is working and inducible promoter is succesfully induced from AI-2.

Applications of BBa_K1202110

User Reviews

UNIQ3424d75a8018db1e-partinfo-00000000-QINU UNIQ3424d75a8018db1e-partinfo-00000001-QINU