Team:UNITN-Trento/Protocols
From 2013.igem.org
(Difference between revisions)
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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}} | ||
+ | |||
+ | {{:Team:UNITN-Trento/Templates/Styles/ProtocolSpoiler|Ethylene detection through Micro GC|<html> | ||
+ | <p> | ||
+ | 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 °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 | ||
+ | column (see the tables for colums and method specifications). | ||
+ | </p> | ||
+ | <table class="tn-sp-table"> | ||
+ | <TR> | ||
+ | <th> | ||
+ | Column | ||
+ | </th> | ||
+ | <th> | ||
+ | Lenght | ||
+ | </th> | ||
+ | <th> | ||
+ | Diameter | ||
+ | </th> | ||
+ | </TR> | ||
+ | <tr> | ||
+ | <td> | ||
+ | Mol Sieve 5A Plot | ||
+ | </td> | ||
+ | <td> | ||
+ | 10 m | ||
+ | </td> | ||
+ | <td> | ||
+ | 0.32 mm | ||
+ | </td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td> | ||
+ | Plot U | ||
+ | </td> | ||
+ | <td> | ||
+ | 8 m | ||
+ | </td> | ||
+ | <td> | ||
+ | 0.32 mm | ||
+ | </td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | <table class="tn-sp-table"> | ||
+ | <tr> | ||
+ | <th colspan = '2'> | ||
+ | Method used | ||
+ | </th> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <th> | ||
+ | t sample | ||
+ | </th> | ||
+ | <td> | ||
+ | 50 °C | ||
+ | </td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <th> | ||
+ | t injector | ||
+ | </th> | ||
+ | <td> | ||
+ | 55 °C | ||
+ | </td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <th> | ||
+ | t column Mol Sieve 5A Plot | ||
+ | </th> | ||
+ | <td> | ||
+ | 110 °C | ||
+ | </td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <th> | ||
+ | t column Plot U | ||
+ | </th> | ||
+ | <td> | ||
+ | 70 °C | ||
+ | </td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <th> | ||
+ | p column Mol Sieve 5A Plot | ||
+ | </th> | ||
+ | <td> | ||
+ | 39.16 psi | ||
+ | </td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <th> | ||
+ | p column Plot U | ||
+ | </th> | ||
+ | <td> | ||
+ | 21.76 psi | ||
+ | </td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <th> | ||
+ | t injection | ||
+ | </th> | ||
+ | <td> | ||
+ | 40 us | ||
+ | </td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <th> | ||
+ | t analysis | ||
+ | </th> | ||
+ | <td> | ||
+ | 95 s | ||
+ | </td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | A measure was then taken.<br/> | ||
+ | In order to estimate how much gas was taken for each measurement using the settings | ||
+ | described above, a mass flow meter was connected to the micro GC. | ||
+ | During a measurment, a flow of 3 (± 0.15) ml / min was registered. Due to the fact that a | ||
+ | measurment lasts 10 s, the withdrawn volume was 0.5 ml. | ||
+ | </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