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Team:British Columbia/Notebook/Week 15
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Joe expressed the proteins and spun down the cell pellets on the weekend. Now, Daisy and Joe are going to be doing the protein purification, in vitro assay and the GCMS sample preparation for alpha-pinene and beta-pinene. Daisy and Joe took 1-2 grams of cell pellet and sonicated the cell pellet with cell lysis buffer. The samples were spun down and the supernatant was collected. The supernatant was put through a gravity nickel column. It was washed several times and then eluted with His-column elution buffer. The enzymes were centrifuged onto a membrane and then re-eluted with the correct enzyme assay buffer, to allow the enzymatic activity to occur. The enzymes were incubated with geranyl diphosphate (GPP) and the samples were prepared. | Joe expressed the proteins and spun down the cell pellets on the weekend. Now, Daisy and Joe are going to be doing the protein purification, in vitro assay and the GCMS sample preparation for alpha-pinene and beta-pinene. Daisy and Joe took 1-2 grams of cell pellet and sonicated the cell pellet with cell lysis buffer. The samples were spun down and the supernatant was collected. The supernatant was put through a gravity nickel column. It was washed several times and then eluted with His-column elution buffer. The enzymes were centrifuged onto a membrane and then re-eluted with the correct enzyme assay buffer, to allow the enzymatic activity to occur. The enzymes were incubated with geranyl diphosphate (GPP) and the samples were prepared. | ||
| - | Originally, the rate of the gravity filtration of the nickel column was very slow and would mean that we were going to be there for a very long time. According to Derek (alina's lab mate), we would be in the lab until 11:30 pm if we were to do the | + | Originally, the rate of the gravity filtration of the nickel column was very slow and would mean that we were going to be there for a very long time. According to Derek (alina's lab mate), we would be in the lab until 11:30 pm if we were to do the purification step. We did stay in the lab until 12 am. |
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===Team Social=== | ===Team Social=== | ||
Joe, Daisy, Alina, Jacob, Rafael, Marianne went for a team social! | Joe, Daisy, Alina, Jacob, Rafael, Marianne went for a team social! | ||
Revision as of 04:01, 14 September 2011
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Protein Purification of Alpha and Beta-Pinene Synthases
Joe expressed the proteins and spun down the cell pellets on the weekend. Now, Daisy and Joe are going to be doing the protein purification, in vitro assay and the GCMS sample preparation for alpha-pinene and beta-pinene. Daisy and Joe took 1-2 grams of cell pellet and sonicated the cell pellet with cell lysis buffer. The samples were spun down and the supernatant was collected. The supernatant was put through a gravity nickel column. It was washed several times and then eluted with His-column elution buffer. The enzymes were centrifuged onto a membrane and then re-eluted with the correct enzyme assay buffer, to allow the enzymatic activity to occur. The enzymes were incubated with geranyl diphosphate (GPP) and the samples were prepared.
Originally, the rate of the gravity filtration of the nickel column was very slow and would mean that we were going to be there for a very long time. According to Derek (alina's lab mate), we would be in the lab until 11:30 pm if we were to do the purification step. We did stay in the lab until 12 am.
Team Social
Joe, Daisy, Alina, Jacob, Rafael, Marianne went for a team social!
