User:Puja Mody/Notebook/Chem 571: Gold Nanoparticles/2012/10/24: Difference between revisions
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* ADA protein was expressed in the same manner as on [[User:Puja Mody/Notebook/Chem 571: Gold Nanoparticles/2012/09/19| 2012/09/19]] | * ADA protein was expressed in the same manner as on [[User:Puja Mody/Notebook/Chem 571: Gold Nanoparticles/2012/09/19| 2012/09/19]] | ||
** This morning, Michael came in around 9am to centrifuge the starter culture media and resuspended the pellets in 4mL of fresh LB. These resuspended cells were then redivided into the expression culture media which was then inoculated in the shaker at 160rpm and 37°C. | ** This morning, Michael came in around 9am to centrifuge the starter culture media and resuspended the pellets in 4mL of fresh LB. These resuspended cells were then redivided into the expression culture media which was then inoculated in the shaker at 160rpm and 37°C. This was completed around 10am. | ||
** At approximately 1pm that same day, the IPTG was added into each of the flasks | |||
** 5mL of .4M IPTG solution was made and 1mL was placed into each of the 4 fernbach flasks. | ** 5mL of .4M IPTG solution was made and 1mL was placed into each of the 4 fernbach flasks. | ||
** .005L x.4M= .002 moles x 238.31g/mol= .477g IPTG in 5mL. Actual= .4754g in 5mL of water. | ** .005L x.4M= .002 moles x 238.31g/mol= .477g IPTG in 5mL. Actual= .4754g in 5mL of water. | ||
* The 4 flasks were then shaken for | * The 4 flasks were then shaken for 3 additional hours (until 4pm). After which the cells were spinned down in the centrifuge at 4500rpm for 15 minutes. Two rounds of centrifugation occurred because also the media could not fit into the centrifuge jars at once. | ||
*Supernatant was poured out and 30mL of binding buffer was used to resusped the cells. The flask of buffer and cells was then placed in a -80°C freezer for 1 week. | |||
* For information regarding the Lysozyme please see either [[User:Michael F. Nagle/Notebook/Chem 571/2012/10/24| Mikes notebook]] or [[User:Mary Mendoza/Notebook/CHEM 571 Experimental Biological Chemistry I/2012/10/24| Marys notebook]]. The lysozyme did create purple gold nanoparticle fibers upon removal from the oven. | * For information regarding the Lysozyme please see either [[User:Michael F. Nagle/Notebook/Chem 571/2012/10/24| Mikes notebook]] or [[User:Mary Mendoza/Notebook/CHEM 571 Experimental Biological Chemistry I/2012/10/24| Marys notebook]]. The lysozyme did create purple gold nanoparticle fibers upon removal from the oven. |
Revision as of 11:58, 24 November 2012
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Transformation, protein expression, lysozyme Uv-VisGoals
Procedure
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