Team:Chiba/Sender experiments/Senders(XL10Gold) T9002(JW1908)

From 2008.igem.org

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(Reaction temparature:30°C)
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__NOTOC__
__NOTOC__
==Results==
==Results==
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===Reaction temparature:37°C===
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===Reaction temparature:37°C===
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*Sender culture:500μL,Receiver:500μL
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====Sender culture:500μL,Receiver:500μL====
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[[Image:Chiba_communication_XL10Gol_37_RS1_01.gif‎|thumb|left|'''Fig.1'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37&deg;C]]
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[[Image:Chiba_communication_XL10Gol_37_RS1_01.gif‎|thumb|left|'''Fig.1'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37&deg;C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
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[[Image:Chiba_talks_XL10G_37_RS1_02.gif‎|thumb|left|'''Fig.2'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37&deg;C]]
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[[Image:Chiba_talks_XL10G_37_RS1_02.gif‎|thumb|left|'''Fig.2'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37&deg;C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
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[[Image:Chiba_talks_XL10G_37_RS1_03.gif‎|thumb|left|'''Fig.3'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37&deg;C]]
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[[Image:Chiba_talks_XL10G_37_RS1_03.gif‎|thumb|left|'''Fig.3'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37&deg;C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
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*result
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*Fig. 1
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Cultures containing cells transformed with LuxI gene([http://partsregistry.org/Part:BBa_K084012 BBa_K084012])and cells transformed with RhlI gene([http://partsregistry.org/Part:BBa_K084008 BBa_K084008]) express gfp and fluorescence intensity was reached to 500.Cultures containing cells transformed with LasI gene([http://partsregistry.org/Part:BBa_K084007 BBa_K084007]) weakly expressed gfp,and fluorescence intensity was not reached to a significant level.
+
*results
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*Discussion:Maximum fluorescence intensity was vary,however,time before gfp expression doesn't differ.
+
**Cultures containing cells transformed with LuxI gene([http://partsregistry.org/Part:BBa_K084012 BBa_K084012])and cells transformed with RhlI gene([http://partsregistry.org/Part:BBa_K084008 BBa_K084008]) express gfp and fluorescence intensity was reached to 500.Cultures containing cells transformed with LasI gene([http://partsregistry.org/Part:BBa_K084007 BBa_K084007]) weakly expressed gfp,and fluorescence intensity was not reached to a significant level.
-
 
+
*Discussion
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*We concluded that LasI gene([http://partsregistry.org/Part:BBa_K084007 BBa_K084007]) was not work well at this condition.
+
#Maximum fluorescence intensity was vary,however,time before gfp expression doesn't differ.
 +
#We concluded that LasI gene([http://partsregistry.org/Part:BBa_K084007 BBa_K084007]) was not work well at this condition.
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middle
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*Fig. 2
*Results
*Results
Fluorescence intensity of the culture containing cells transformed with LuxI gene([http://partsregistry.org/Part:BBa_K084012 BBa_K084012]) was lower than that of the cultures containing the cells transformed with LVA LuxI-LVA gene([http://partsregistry.org/Part:BBa_K084014 BBa_K084014]).The difference of maximum fluorescence intesity between the two cultures was 200.
Fluorescence intensity of the culture containing cells transformed with LuxI gene([http://partsregistry.org/Part:BBa_K084012 BBa_K084012]) was lower than that of the cultures containing the cells transformed with LVA LuxI-LVA gene([http://partsregistry.org/Part:BBa_K084014 BBa_K084014]).The difference of maximum fluorescence intesity between the two cultures was 200.
*Discussion
*Discussion
 +
**The rate of AHL synthesis decreased by the degradation of autoinducer synthase,however,the length of time before gfp expression is same.
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*The rate of AHL synthesis decreased by the degradation of autoinducer synthase,however,the length of time before gfp expression is same.
 
-
 
+
*Fig. 3
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*right
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*Results
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*Result
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Cultures containing cells transformed with RhlI gene([http://partsregistry.org/Part:BBa_K084008 BBa_K084008]) and RhlI+LVA([http://partsregistry.org/Part:BBa_K084009 BBa_K084009]) draw the same transfer curve.
Cultures containing cells transformed with RhlI gene([http://partsregistry.org/Part:BBa_K084008 BBa_K084008]) and RhlI+LVA([http://partsregistry.org/Part:BBa_K084009 BBa_K084009]) draw the same transfer curve.
*Discussion
*Discussion
 +
**We concluded that there was no effect of LVA-tag on time before gfp expression. 
-
*We concluded that there was no effect of LVA-tag on time before gfp expression. 
 
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[[Image:Chiba_talks_XL10Gold_37_RS1_0917_01.gif|thumb|left|'''Fig.4'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C]]
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[[Image:Chiba_talks_XL10Gold_37_RS1_0917_01.gif|thumb|left|'''Fig.4'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
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[[Image:Chiba_talks_XL10Gold_37_RS1_0917_02.gif|thumb|left|'''Fig.5'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C]]
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[[Image:Chiba_talks_XL10Gold_37_RS1_0917_02.gif|thumb|left|'''Fig.5'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
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 +
 +
====Sender culture:100&mu;L,Receiver:1000&mu;L====
 +
[[Image:Chiba_talks_XL10Gold_37_RS1_01.gif|thumb|left|'''Fig. 6'''  
 +
<br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37&deg;C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
 +
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 +
 +
====Sender culture:10&mu;L,Receiver:1000&mu;L====
===Reaction temparature:30°C===
===Reaction temparature:30°C===
*Sender culture:500&mu;L, Receiver culture:500&mu;L
*Sender culture:500&mu;L, Receiver culture:500&mu;L
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[[Image:Chiba_talks_XL10G_30-1_RS1_01.gif‎‎|thumb|left|'''Fig.6'''  <br>E.coli strain,BBa_K084007:XL10Gold,BBa_T9002:JW1908,30°C]]
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[[Image:Chiba_talks_XL10G_30-1_RS1_01.gif‎‎|thumb|left|'''Fig.7'''  <br>E.coli strain,BBa_K084007:XL10Gold,BBa_T9002:JW1908,30°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
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[[Image:Chiba_talks_XL10G_30-1_RS1_02.gif‎‎‎|thumb|left|'''Fig.7'''  <br>E.coli strain,BBa_K084007:XL10Gold,BBa_T9002:JW1908,30°C]]
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[[Image:Chiba_talks_XL10G_30-1_RS1_02.gif‎‎‎|thumb|left|'''Fig.8'''  <br>E.coli strain,BBa_K084007:XL10Gold,BBa_T9002:JW1908,30°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
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<br clear="all">
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*
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*Fig. 7
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**Result  
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*Result  
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***蛍光強度はRhl,LuxI+LVA,LasIの順に高かった。
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#The highest maximum fluorescence intensity was caused by RhlI gene,followed by LuxI(with LVA) and LasI gene.
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***LuxI,RhlIとLasIは蛍光強度200に達するまでの時間差は約2時間だった
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#Fluorescence intensity of the two culture containing cells transformed with LuxI gene and RhlI gene amount to 200 for 2 hour as fast as  LasI gene.
-
**考察
+
*Discussion
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***30&deg;CではRhlIの活性のほうがLuxIより高い。もしくは、LVAの効果が出ている。その場合、右のグラフ
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#Maximum fluorescence intensity caused by RhlI gene was higher than that caused by LuxI gene.There are two reasons:
-
   のLVA自体に問題があると考えられる。
+
#RhlI was more active at this experimental condition.
 +
#LuxI was well degraded by protease.
 +
#We concluded that LVA-tag effective in this condition.
-
*
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*Fig. 8
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**Result  
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*Result  
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***RhlI+LVAとRhlIによるAHL合成量を蛍光強度で比較した場合、値はほぼ同じだった
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**Comparing RhlI with  Rhl+LVAtag, fluoroscence intensity were almost same.
-
**考察
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*Discussion
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*** LVAが働いていないか、働いていたとしてもこの条件だとAHLシンターゼの合成速度のほうがLVAによるLuxIの分解速度よりも
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**In this condition, production efficiency of LuxI protein higher than of LVAtag.
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  大きいと考えられる
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-
===Reaction temparature:25°C===
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===Reaction temparature:25&deg;C===
====Sender culture:500μL,Receiver culture:500μL====
====Sender culture:500μL,Receiver culture:500μL====
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[[Image:Chiba_talks_XL10Gold_25_RS1_01.gif|thumb|left|'''Fig.8'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,25°C,Receiver cells/Sender cells = 1.]]
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[[Image:Chiba_talks_XL10Gold_25_RS1_01.gif|thumb|left|'''Fig.9'''  <br>E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,25&deg;C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
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[[Image:Chiba_talks_XL10Gold_25_RS1_02.gif|thumb|left|'''Fig.9''' <br> E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,25°C,Receiver cells/Sender cells = 1.]]
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[[Image:Chiba_talks_XL10Gold_25_RS1_02.gif|thumb|left|'''Fig.10''' <br> E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,25&ded;C.All measurements are averages from three replicate cultures with error bars representing standard deviations.]]
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<br clear=all>
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*Left:
+
*Fig.9
-
**result
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*results
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***30&deg;C,37&deg;Cの実験時に比べて蛍光強度が極端に低く,ネガコンと差がほとんどなかった.
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#The value of fluoroscence intensity by LuxI, RhlI, LasI gene at 25&deg;c  were almost the same as negative control at 30 and 37&deg;c.  
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***LuxI,RhlIに比べてLasIにおける最終形高強度は約1/2と低いものであった。
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#The last value of fluoroscence intensity by LasI gene was a half of LuxI and RhlI.
-
**Discussion
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*Discussion
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***蛍光強度が極端に下がったのは30°C,37°Cで行った実験と比べ温度が低いため活性が落ちたためと考えられる。
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#Low temperature of 25&deg;c caused decrease of production efficiency of AHL synthase.
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***またそれだけではなく静置して行ったためSenderとReceiverがよく混ざらず、その結果GFPの発現量が大幅に減少した可能性も考えられる。
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#Static culture keep sender and receiver from shaking, in the end , expression of GFP decreased.
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*Right:
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*Fig.10
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**Result
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*Result
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***The fluoroscence intensity by RhlI with LVAtag is less than RhlI without LVAtag.
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**The fluoroscence intensity by RhlI with LVAtag is less than RhlI without LVAtag.
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**Discussion  
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*Discussion  
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***LVAtag is effective in this condition.
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**LVAtag is effective in this condition.
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[[Team:Chiba/Project/Experiments:Sender_Crosstalk||Back to Sender experiment and result]]
[[Team:Chiba/Project/Experiments:Sender_Crosstalk||Back to Sender experiment and result]]

Latest revision as of 09:47, 30 October 2008

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


Results

Reaction temparature:37°C

Sender culture:500μL,Receiver:500μL

Fig.1  
E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.
Fig.2  
E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.
Fig.3  
E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.


  • Fig. 1
  • results
    • Cultures containing cells transformed with LuxI gene([http://partsregistry.org/Part:BBa_K084012 BBa_K084012])and cells transformed with RhlI gene([http://partsregistry.org/Part:BBa_K084008 BBa_K084008]) express gfp and fluorescence intensity was reached to 500.Cultures containing cells transformed with LasI gene([http://partsregistry.org/Part:BBa_K084007 BBa_K084007]) weakly expressed gfp,and fluorescence intensity was not reached to a significant level.
  • Discussion
  1. Maximum fluorescence intensity was vary,however,time before gfp expression doesn't differ.
  2. We concluded that LasI gene([http://partsregistry.org/Part:BBa_K084007 BBa_K084007]) was not work well at this condition.
  • Fig. 2
  • Results

Fluorescence intensity of the culture containing cells transformed with LuxI gene([http://partsregistry.org/Part:BBa_K084012 BBa_K084012]) was lower than that of the cultures containing the cells transformed with LVA LuxI-LVA gene([http://partsregistry.org/Part:BBa_K084014 BBa_K084014]).The difference of maximum fluorescence intesity between the two cultures was 200.

  • Discussion
    • The rate of AHL synthesis decreased by the degradation of autoinducer synthase,however,the length of time before gfp expression is same.


  • Fig. 3
  • Results

Cultures containing cells transformed with RhlI gene([http://partsregistry.org/Part:BBa_K084008 BBa_K084008]) and RhlI+LVA([http://partsregistry.org/Part:BBa_K084009 BBa_K084009]) draw the same transfer curve.

  • Discussion
    • We concluded that there was no effect of LVA-tag on time before gfp expression.



Fig.4  
E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.
Fig.5  
E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.



Sender culture:100μL,Receiver:1000μL

Fig. 6  
E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,37°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.


Sender culture:10μL,Receiver:1000μL

Reaction temparature:30°C

  • Sender culture:500μL, Receiver culture:500μL
Fig.7  
E.coli strain,BBa_K084007:XL10Gold,BBa_T9002:JW1908,30°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.
Fig.8  
E.coli strain,BBa_K084007:XL10Gold,BBa_T9002:JW1908,30°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.


  • Fig. 7
  • Result  
  1. The highest maximum fluorescence intensity was caused by RhlI gene,followed by LuxI(with LVA) and LasI gene.
  2. Fluorescence intensity of the two culture containing cells transformed with LuxI gene and RhlI gene amount to 200 for 2 hour as fast as LasI gene.
  • Discussion
  1. Maximum fluorescence intensity caused by RhlI gene was higher than that caused by LuxI gene.There are two reasons:
  2. RhlI was more active at this experimental condition.
  3. LuxI was well degraded by protease.
  4. We concluded that LVA-tag effective in this condition.


  • Fig. 8
  • Result
    • Comparing RhlI with Rhl+LVAtag, fluoroscence intensity were almost same.
  • Discussion
    • In this condition, production efficiency of LuxI protein higher than of LVAtag.

Reaction temparature:25°C

Sender culture:500μL,Receiver culture:500μL

Fig.9
E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,25°C.All measurements are averages from three replicate cultures with error bars representing standard deviations.
Fig.10
 E.coli strain,Senders:XL10Gold,BBa_T9002:JW1908,25&ded;C.All measurements are averages from three replicate cultures with error bars representing standard deviations.


  • Fig.9
  • results
  1. The value of fluoroscence intensity by LuxI, RhlI, LasI gene at 25°c were almost the same as negative control at 30 and 37°c.
  2. The last value of fluoroscence intensity by LasI gene was a half of LuxI and RhlI.
  • Discussion
  1. Low temperature of 25°c caused decrease of production efficiency of AHL synthase.
  2. Static culture keep sender and receiver from shaking, in the end , expression of GFP decreased.
  • Fig.10
  • Result
    • The fluoroscence intensity by RhlI with LVAtag is less than RhlI without LVAtag.
  • Discussion
    • LVAtag is effective in this condition.


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