Difference between revisions of "Team:Gifu/InterLab"

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<center><font size="20" face="Century">Inter Lab Study</font></center><br><br>
 
<center><font size="20" face="Century">Inter Lab Study</font></center><br><br>
  
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<font size="6" face="Arimo">Equipment</font>  
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<hr width="100%" ><br>
 
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<ul>
 
<ul>
 
<li> <font size="5" face="Arimo"> BioSHAKER TA-12R<br></font> </li>
 
<li> <font size="5" face="Arimo"> BioSHAKER TA-12R<br></font> </li>
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use for shaking culture</font>
 
use for shaking culture</font>
 
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<li> <font size="5" face="Arimo"> Safire ART-NR:F129013<br></font> </li>
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<li> <font size="5" face="century"> Safire ART-NR:F129013<br></font> </li>
 
<font size="4" face="Arimo">
 
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use for measurement of absorbance</font>
 
use for measurement of absorbance</font>
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<font size="6" face="Arimo">Method of Measurement</font>  
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<font size="6" face="century">Method of Measurement</font>  
 
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<p>&nbsp;&nbsp;We constructed 5 plasmids [positive control, negative control, device1~3] by using BioBricks and then transformed to <i>E. coli</i> K-12 JM109. We checked them by colony direct PCR and plasmids sequence. We cultivated correct bacterias by shaking until OD660:0.5. Cultivated mediums were　transferred to 96-well plate by 100mL. We read the absorbance.</p>
 
<p>&nbsp;&nbsp;We constructed 5 plasmids [positive control, negative control, device1~3] by using BioBricks and then transformed to <i>E. coli</i> K-12 JM109. We checked them by colony direct PCR and plasmids sequence. We cultivated correct bacterias by shaking until OD660:0.5. Cultivated mediums were　transferred to 96-well plate by 100mL. We read the absorbance.</p>
 
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<font size="6" face="Arimo">Result</font>  
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<li> <font size="5" face="Arimo"> Definition<br></font> </li>
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<li> <font size="5" face="century"> Definition<br></font> </li>
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<p>&nbsp;&nbsp;We expressed absorbance as relative value for　positive control. <br>This expression is following:<br></p></font>  
 
<p>&nbsp;&nbsp;We expressed absorbance as relative value for　positive control. <br>This expression is following:<br></p></font>  
 
<img src="https://static.igem.org/mediawiki/2015/8/85/Siki-gifu.png" border="0" width="211" height="57"><br>
 
<img src="https://static.igem.org/mediawiki/2015/8/85/Siki-gifu.png" border="0" width="211" height="57"><br>
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<p>Relative absorbance values for positive control are shown below. <br></p>
 
<p>Relative absorbance values for positive control are shown below. <br></p>
 
<img src="https://static.igem.org/mediawiki/2015/e/e4/GURAHU-gifu.png" border="0" width="572" height="364"> <br><br>
 
<img src="https://static.igem.org/mediawiki/2015/e/e4/GURAHU-gifu.png" border="0" width="572" height="364"> <br><br>
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<p>&nbsp;&nbsp;The value of device1 is much higher than that of device2 and 3. This is why each device is different from promoter sequence, especially -10 and -35 region.<br><br></p>
 
<p>&nbsp;&nbsp;The value of device1 is much higher than that of device2 and 3. This is why each device is different from promoter sequence, especially -10 and -35 region.<br><br></p>
 
<img src="https://static.igem.org/mediawiki/2015/b/b4/Konnsennsasu2.png" border="0" width="413" height="105"> <br><br>
 
<img src="https://static.igem.org/mediawiki/2015/b/b4/Konnsennsasu2.png" border="0" width="413" height="105"> <br><br>

Revision as of 04:16, 31 August 2015


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Inter Lab Study


Equipment

  • BioSHAKER TA-12R
    • use for shaking culture



  • Safire ART-NR:F129013
    • use for measurement of absorbance




Method of Measurement

      We constructed 5 plasmids [positive control, negative control, device1~3] by using BioBricks and then transformed to E. coli K-12 JM109. We checked them by colony direct PCR and plasmids sequence. We cultivated correct bacterias by shaking until OD660:0.5. Cultivated mediums were　transferred to 96-well plate by 100mL. We read the absorbance.


      Setting of plate reader is following:



Result

  • Definition

    •   We expressed absorbance as relative value for　positive control.
    This expression is following:
      Sample : Absorbance in sample
      NCav : Average of absorbance in negative control
      PCav : Average of absorbance in positive control



  • Value

    • Relative absorbance values for positive control are shown below.



    FIG1 Relative absorbance values
    20A : Device1 (J23101 + I13504)
    22A : Device2 (J23106 + I13504)
    22K : Device3 (J23117 + I13504)




Discussion

      The value of device1 is much higher than that of device2 and 3. This is why each device is different from promoter sequence, especially -10 and -35 region.



    Promoter sequence in device1 resembles consensus sequence. On the other hand, device2 and 3 is not similar than device1. This is how each GFP expression is different.









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