User:Paul Rothenberg/Notebook/Pauls Notebook/2014/06/16: Difference between revisions
From OpenWetWare
| Line 8: | Line 8: | ||
==Restart!== | ==Restart!== | ||
After lyophilizing the Mn Mb last week, we found that there was only 2 mg total produced (out of 200 mg of protein!). We restarted, so we synthesized more ApoMb. We doubled the volume of the Tris buffer to 8 mL with the same concentration, and only used 100 mg of protein, not 200 mg. We are going to be more cautious and ensure that the protein is fully in solution before performing extraction. We brought the pH down to 2.3 with 40 µL of 1 N HCl. | After lyophilizing the Mn Mb last week, we found that there was only 2 mg total produced (out of 200 mg of protein!). We restarted, so we synthesized more ApoMb. We doubled the volume of the Tris buffer to 8 mL with the same concentration, and only used 100 mg of protein, not 200 mg. We are going to be more cautious and ensure that the protein is fully in solution before performing extraction. We then centrifuged the solution at 4,000 rpm for 30 minutes at 25°C before the extraction. | ||
We brought the pH down to 2.3 with 40 µL of 1 N HCl. | |||
We also ran a UV-VIS spectrum for both layers. | We also ran a UV-VIS spectrum for both layers. | ||
Revision as of 08:09, 16 June 2014
 Project name
|
<html><img src="/images/9/94/Report.png" border="0" /></html> Main project page <html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html> </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html> |
Restart!After lyophilizing the Mn Mb last week, we found that there was only 2 mg total produced (out of 200 mg of protein!). We restarted, so we synthesized more ApoMb. We doubled the volume of the Tris buffer to 8 mL with the same concentration, and only used 100 mg of protein, not 200 mg. We are going to be more cautious and ensure that the protein is fully in solution before performing extraction. We then centrifuged the solution at 4,000 rpm for 30 minutes at 25°C before the extraction. We brought the pH down to 2.3 with 40 µL of 1 N HCl. We also ran a UV-VIS spectrum for both layers. Once the extraction was complete, we placed it in the same Na2CO3 buffer for overnight dialysis.
| |
