Team:Chiba/Calendar-Home/16 October 2008

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[[Team:Chiba/Calendar-Home/15 October 2008|15 October 2008 <]]|[[Team:Chiba/Calendar-Home/17 October 2008|> 17 October 2008]]
[[Team:Chiba/Calendar-Home/15 October 2008|15 October 2008 <]]|[[Team:Chiba/Calendar-Home/17 October 2008|> 17 October 2008]]
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==Laboratory work==
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=== Team:Demo-Is ===
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#Pre-culture
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##Picked and cultured the following glycerol stocks in 2mL of LB:
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###LB-Amp, [http://partsregistry.org/Part:BBa_T9002 BBa_T9002], (JW1908)
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###LB-Amp+0.2%Glu, [http://partsregistry.org/Part:BBa_K084012 BBa_K084012]([http://partsregistry.org/Part:BBa_J04500 plac+rbs]+[http://partsregistry.org/wiki/index.php?title=Part:BBa_C0161 LuxI(no LVA)]), (XL10G)
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###LB-Amp+0.2%Glu, [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]([http://partsregistry.org/Part:BBa_J04500 plac+rbs]+[http://partsregistry.org/wiki/index.php?title=Part:BBa_C0178 LasI(no LVA)]), (XL10G)
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##Cultured at 37&deg;C for 12h.
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#Culture
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##Added 6.25% each of the pre-cultures to new LB medium.
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###LB-Amp, [http://partsregistry.org/Part:BBa_T9002 BBa_T9002]
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###LB-Amp+0.2%Glu, [http://partsregistry.org/Part:BBa_K084012 BBa_K084012]([http://partsregistry.org/Part:BBa_J04500 plac+rbs]+[http://partsregistry.org/wiki/index.php?title=Part:BBa_C0161 LuxI(no LVA)]), [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]([http://partsregistry.org/Part:BBa_J04500 plac+rbs]+[http://partsregistry.org/wiki/index.php?title=Part:BBa_C0178 LasI(no LVA)])
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##Cultured at 37&deg;C for 4~5h
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#Wash
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##Transfer 10mL each of the culture to 50mL centrifuge tubes.
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##Centrifuged for 6min at 3600rpm,20&deg;C and discarded the supernatant.
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##Added LB-Amp to each centrifuge tube:
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###10mL to the tube that contains [http://partsregistry.org/Part:BBa_T9002 BBa_T9002]
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###5mL to the tube that contains [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]
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##Centrifuged for 6min, 3600rpm at 20&deg;C the tube containing [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007] and discarded　the supernatant.
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##10mL to the tube that contains [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]
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##Centrifuged for 6min, 3600rpm at 20&deg;C the tube containing [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007] and discarded　the supernatant.
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##5mL to the tube that contains [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]
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#Mix
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##Mixed the sender cells [http://partsregistry.org/Part:BBa_K084012 BBa_K084012] and [http://partsregistry.org/Part:BBa_K084007 BBa_K084007] both with [http://partsregistry.org/Part:BBa_T9002 BBa_T9002] at a 1:1 ratio.
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##Added 100&mu;L each to a 96-well shallow plate (as shown in the figure).
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###Green part is [http://partsregistry.org/Part:BBa_K084012 BBa_K084012]:[http://partsregistry.org/Part:BBa_T9002 BBa_T9002]=1:1
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###Red part is [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]:[http://partsregistry.org/Part:BBa_T9002 BBa_T9002]=1:1
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###Uncolored part is [http://partsregistry.org/Part:BBa_T9002 BBa_T9002] alone.
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#Culture and observe results
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====Results====
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[[Image:Team-Chiba-demo-mihon.gif|200px]] Green region: sender=LuxI,
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Red circular region: sender=Las I.
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<gallery>
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Image:Team-Chiba-demo-1.JPG|0 h
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Image:Team-Chiba-demo-2.JPG|4 h <br>(Lux I GFP detected)
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Image:Team-Chiba-demo-3.JPG|8 h <br>(Lux I GFP and Las I GFP detected)
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</gallery>
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LuxI GFP is detected at 4h following mixing while LasI GFP is detected
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after 8h, thus successfully demonstrating time-delay depending on the
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sender used.

Latest revision as of 00:50, 30 October 2008

>Home | Notebook

15 October 2008 <|> 17 October 2008

Laboratory work

Team:Demo-Is

  1. Pre-culture
    1. Picked and cultured the following glycerol stocks in 2mL of LB:
      1. LB-Amp, [http://partsregistry.org/Part:BBa_T9002 BBa_T9002], (JW1908)
      2. LB-Amp+0.2%Glu, [http://partsregistry.org/Part:BBa_K084012 BBa_K084012]([http://partsregistry.org/Part:BBa_J04500 plac+rbs]+[http://partsregistry.org/wiki/index.php?title=Part:BBa_C0161 LuxI(no LVA)]), (XL10G)
      3. LB-Amp+0.2%Glu, [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]([http://partsregistry.org/Part:BBa_J04500 plac+rbs]+[http://partsregistry.org/wiki/index.php?title=Part:BBa_C0178 LasI(no LVA)]), (XL10G)
    2. Cultured at 37°C for 12h.
  2. Culture
    1. Added 6.25% each of the pre-cultures to new LB medium.
      1. LB-Amp, [http://partsregistry.org/Part:BBa_T9002 BBa_T9002]
      2. LB-Amp+0.2%Glu, [http://partsregistry.org/Part:BBa_K084012 BBa_K084012]([http://partsregistry.org/Part:BBa_J04500 plac+rbs]+[http://partsregistry.org/wiki/index.php?title=Part:BBa_C0161 LuxI(no LVA)]), [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]([http://partsregistry.org/Part:BBa_J04500 plac+rbs]+[http://partsregistry.org/wiki/index.php?title=Part:BBa_C0178 LasI(no LVA)])
    2. Cultured at 37°C for 4~5h
  3. Wash
    1. Transfer 10mL each of the culture to 50mL centrifuge tubes.
    2. Centrifuged for 6min at 3600rpm,20°C and discarded the supernatant.
    3. Added LB-Amp to each centrifuge tube:
      1. 10mL to the tube that contains [http://partsregistry.org/Part:BBa_T9002 BBa_T9002]
      2. 5mL to the tube that contains [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]
    4. Centrifuged for 6min, 3600rpm at 20°C the tube containing [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007] and discarded　the supernatant.
    5. 10mL to the tube that contains [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]
    6. Centrifuged for 6min, 3600rpm at 20°C the tube containing [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007] and discarded　the supernatant.
    7. 5mL to the tube that contains [http://partsregistry.org/Part:BBa_K084012 BBa_K084012], [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]
  4. Mix
    1. Mixed the sender cells [http://partsregistry.org/Part:BBa_K084012 BBa_K084012] and [http://partsregistry.org/Part:BBa_K084007 BBa_K084007] both with [http://partsregistry.org/Part:BBa_T9002 BBa_T9002] at a 1:1 ratio.
    2. Added 100μL each to a 96-well shallow plate (as shown in the figure).
      1. Green part is [http://partsregistry.org/Part:BBa_K084012 BBa_K084012]:[http://partsregistry.org/Part:BBa_T9002 BBa_T9002]=1:1
      2. Red part is [http://partsregistry.org/Part:BBa_K084007 BBa_K084007]:[http://partsregistry.org/Part:BBa_T9002 BBa_T9002]=1:1
      3. Uncolored part is [http://partsregistry.org/Part:BBa_T9002 BBa_T9002] alone.
  5. Culture and observe results

Results

Team-Chiba-demo-mihon.gif Green region: sender=LuxI, Red circular region: sender=Las I.

0 h

4 h
(Lux I GFP detected)

8 h
(Lux I GFP and Las I GFP detected)


LuxI GFP is detected at 4h following mixing while LasI GFP is detected after 8h, thus successfully demonstrating time-delay depending on the sender used.

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