Team:UNITN-Trento/Protocols

From 2013.igem.org

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Then run the PCR samples on a 1% agarose gel to verify the success of the reactions (each sample is prepared with 8µl of PCR reaction and 2µl of 6X loading die; 1kb ladder is good).
Then run the PCR samples on a 1% agarose gel to verify the success of the reactions (each sample is prepared with 8µl of PCR reaction and 2µl of 6X loading die; 1kb ladder is good).
</html>|pSB1C3-linearization-PCR}}
</html>|pSB1C3-linearization-PCR}}
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{{:Team:UNITN-Trento/Templates/Styles/ProtocolSpoiler|Ethylene detection through Micro GC|<html>
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<p>
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An overnight culture was diluited 1:100 and grown until O.D.600 reached 0.5. After that, the culture was induced with 5 mM arabinose and placed with a stirrer in a sealed vial (V = 15 ml) with a pierceable septum. The sample was kept for about 4 hours at 37 &deg;C in thermoshaker. After that the sample was connected to the Micro Gas Chromatograph Agilent 3000A endowed with two colums: a Mol Sieve 5A Plot and a Plot U
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column (see the tables for colums and method specifications).
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</p>
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<table class="tn-sp-table">
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<TR>
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<th>
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Column
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</th>
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<th>
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Lenght
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</th>
 +
<th>
 +
Diameter
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</th>
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</TR>
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<tr>
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<td>
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Mol Sieve 5A Plot
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</td>
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<td>
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10 m
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</td>
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<td>
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0.32 mm
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</td>
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</tr>
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<tr>
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<td>
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Plot U
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</td>
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<td>
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8 m
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</td>
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<td>
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0.32 mm
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</td>
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</tr>
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</table>
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<table class="tn-sp-table">
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<tr>
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<th colspan = '2'>
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Method used
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</th>
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</tr>
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<tr>
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<th>
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t sample
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</th>
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<td>
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50 &deg;C
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</td>
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</tr>
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<tr>
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<th>
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t injector
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</th>
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<td>
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55 &deg;C
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</td>
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</tr>
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<tr>
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<th>
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t column Mol Sieve 5A Plot
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</th>
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<td>
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110 &deg;C
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</td>
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</tr>
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<tr>
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<th>
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t column Plot U
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</th>
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<td>
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70 &deg;C
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</td>
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</tr>
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<tr>
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<th>
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p column Mol Sieve 5A Plot
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</th>
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<td>
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39.16 psi
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</td>
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</tr>
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<tr>
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<th>
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p column Plot U
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</th>
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<td>
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21.76 psi
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</td>
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</tr>
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<tr>
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<th>
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t injection
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</th>
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<td>
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40 us
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</td>
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</tr>
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<tr>
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<th>
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t analysis
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</th>
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<td>
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95 s
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</td>
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</tr>
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</table>
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A measure was then taken.<br/>
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In order to estimate how much gas was taken for each measurement using the settings
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described above, a mass flow meter was connected to the micro GC.
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During a measurment, a flow of 3 (± 0.15) ml / min was registered. Due to the fact that a
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measurment lasts 10 s, the withdrawn volume was 0.5 ml.
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</html>|ethylene-detection-assay}}
{{:Team:UNITN-Trento/Templates/Styles/ProtocolSpoiler|Kinetic assay for ethylene production through micro GC|<html>
{{:Team:UNITN-Trento/Templates/Styles/ProtocolSpoiler|Kinetic assay for ethylene production through micro GC|<html>

Revision as of 13:15, 20 August 2013


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