Team:Chiba/AiiA Receiver Phase

From 2008.igem.org

(Difference between revisions)
(→AiiA Receiver)
(→AiiA Receiver)
Line 33: Line 33:
**non-AiiA Receiver
**non-AiiA Receiver
[[Image:Non-AiiA-Receiver Chiba.gif]]
[[Image:Non-AiiA-Receiver Chiba.gif]]
 +
===Method===
 +
#Transformed Sender into E.coli strains(BWΔFliC) and Receivers into E.coli strain(BWΔFliC).
 +
#Inoculated them independently in liquid media. Incubated at 37c&deg； 12h.
 +
#Inoculated again at 37c&deg； upto about OD600=2.0
 +
#Washed them.
 +
#Mixed them (Sender:Receiver=1000μl:1000μl).
 +
#Incubated at 30°C.
 +
#Measured intensity of green fluorescence at regular time intervals.
 +
<br clear=all>
===result===
===result===

Revision as of 12:32, 29 October 2008

Chiba-U.gif

Home The Team The Project Parts Submitted to the Registry Reference Notebook Acknowledgements

AiiA Receiver

Fig. AiiA Receiver


Design

オートインデューサー不活性化酵素であるAiiAをレシーバー菌で発現させることにより、レシーバー菌内でのAHL濃度の増加が遅くなるため、AiiAを発現しないレシーバー菌に比べて、GFP発現が遅くなる。（小林）


Fig. AiiA Reaction


Experiment

LuxI-sender Chiba.gif


  • Receivers
    • AiiA Receiver

AiiA-Receiver-genetic-circu Chiba.gif

    • non-AiiA Receiver

Non-AiiA-Receiver Chiba.gif

Method

  1. Transformed Sender into E.coli strains(BWΔFliC) and Receivers into E.coli strain(BWΔFliC).
  2. Inoculated them independently in liquid media. Incubated at 37c&deg； 12h.
  3. Inoculated again at 37c&deg； upto about OD600=2.0
  4. Washed them.
  5. Mixed them (Sender:Receiver=1000μl:1000μl).
  6. Incubated at 30°C.
  7. Measured intensity of green fluorescence at regular time intervals.


result

Fig.Time Delay Test

24時間後の蛍光強度を比較すると最大強度は1/4に下がっている。

transfer curveがシグモイドではなく、時間に比例するようになった

AiiAを発現させると、GFPの発現自体が極端に減ってしまったので、AHL自体を減らしてしまうと発現量の最大値が小さくなってしまう。

conclusion

>Back to the project page

Home The Team The Project Parts Submitted to the Registry Notebook
Retrieved from "http://2008.igem.org/Team:Chiba/AiiA_Receiver_Phase"