Difference between revisions of "Team:UST Beijing/Notebook"
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<li><a href="https://2016.igem.org/Team:UST_Beijing/Demonstrate">Mixed Fermentation</a></li> | <li><a href="https://2016.igem.org/Team:UST_Beijing/Demonstrate">Mixed Fermentation</a></li> | ||
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| − | <li><a href=" | + | <li><a href="https://2016.igem.org/Team:UST_Beijing/AnimalExperiment">Animal Experiment</a></li> |
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Revision as of 12:08, 14 October 2016
Notebook
As early as April of this year, we started to prepare for our project. After the brainstorming, we thought we had a perfect idea, and decided to take action. There have been many failures in the process of experiment, but these failures make the success even more exciting. Here records good memories of these months.
Preparation I: Refer to Literature
April to May
We searched some literature on notoginseng and glucosidase in the database and learned about these papers. We revived E. coli containing β-glucosidase and extracted the plasmid of β-glu. Then the plasmid was cut with NcoI and XhoI in the 37℃ for 1 h.After electrophoresis, we confirmed that plasmid was correct and it can be used in the next experiments.
Preparation II: Liquid fermentation
May to June
The E. coli was centrifuged and the precipitation was immersed with different osmotic pressure buffer to draw β-glu expressed by E. coli. After drawing enzyme, we added different volumes of enzyme solution and pNPG to each well on the 96-well plate and made the bulk equal with extract buffer. OD450 was measured every one hour.
Preparation III: Enzyme activity determination
April to May
E. coli expressed β-glucosidase was fermented in a 3L fermenter. And we collected a small of fermentation sample at 0, 4, 8, 12, 16, 18, 20 and 24 hours since the fermentation start. After centrifugation, β-glu was extracted from the cell. And we tested the activity of the enzyme extracted from different growth stages of E. coli on a 96-well plate with pNPG as the substrate.
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