Team:ITESM-CEM/Interlab
From 2014.igem.org
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<a name="Overview"> <h2>Overview</h2> </a> | <a name="Overview"> <h2>Overview</h2> </a> | ||
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<p>During 2014 iGEM competition, teams were requested to analyse the efficiency of 3 different genetic devices (BioBricks) using GFP as a marker of gene expression. Here, iGEM ITESM CEM team presents the results of this interlab fluorescence measurement study. | <p>During 2014 iGEM competition, teams were requested to analyse the efficiency of 3 different genetic devices (BioBricks) using GFP as a marker of gene expression. Here, iGEM ITESM CEM team presents the results of this interlab fluorescence measurement study. | ||
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<a name="Protocols"> <h2>Protocols</h2> </a> | <a name="Protocols"> <h2>Protocols</h2> </a> | ||
+ | <h1><U>Transformation Protocol</U></h1> | ||
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- | + | All the previously assembled BioBricks were transformed into DH5α competent cells acquired from New England Biolabs (NEB ®). In order to do so, NEB ®’s transformation protocol (3) was used: 50 μl of competent cells were added to microtubes; then, 5 μl of each previously assembled device (DNA concentration between 200 and 300 pg/ml, as determined by spectrophotometry) were pipetted into the tube, which was placed on ice for 30 minutes. Afterwards, the samples were submitted to 30 seconds of a 42°C heat shock; after which they were placed on ice for another 5 minutes. After incubation on ice, 950 μl of SOC medium were added to each mixture. <br>The tubes were placed at 37°C and 250 rpm for 60 minutes. Finally, 200 μl of each sample were plated into warm, solid LB media with 0.1% v/v of antibiotic (kanamycin 15 mg/ml for device 1, and chloramphenicol 35 mg/ml for devices 2, and 3).<br> | |
+ | After 12 hours of incubation at 37°C, a single colony was isolated from each plate and cultured overnight in liquid LB media with the previously stated concentration of antibiotic. A part of these liquid cultures was used for plasmid extraction and isolation; while the rest of them was used to perform relative fluorescence measurements. Both extraction and fluorescence measurement protocols are described later. | ||
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Revision as of 18:30, 25 September 2014
ITESM-CEM | Enzy7-K me |
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