"
Team:Cambridge/Experiments/Protein Purification
From 2011.igem.org
(Difference between revisions)
(→Protein Purification) |
(→Practice) |
||
| Line 7: | Line 7: | ||
*After high copy expression plasmids for his-tagged reflectin were successfully [assembled] and [transformed] in E. coli, cultures were incubated overnight with () arabinose to induce reflectin expression. | *After high copy expression plasmids for his-tagged reflectin were successfully [assembled] and [transformed] in E. coli, cultures were incubated overnight with () arabinose to induce reflectin expression. | ||
*[Buffers] were prepared, and their pH checked and readjusted if necessary on the day of purification. | *[Buffers] were prepared, and their pH checked and readjusted if necessary on the day of purification. | ||
| - | *An [inclusion body prep] was performed with 50ml of overnight culture. | + | *An [inclusion body prep] was performed with 50ml of overnight culture. Reflectin was [purified] from the resulting lysate using a his-trap column. |
| + | *This procedure was repeated using a culture of the same bacteria from a different flask. | ||
| + | |||
| + | ==Results== | ||
| + | Photospectroscopy readings from the eluted solution indicated that we had | ||
Revision as of 10:54, 21 August 2011
Loading...
Protein Purification
Bacteria expressing his-tagged reflectin were lysed, and the protein was purified using a his-trap column and a denaturing protocol in order to solubilise reflectin.
Practice
- After high copy expression plasmids for his-tagged reflectin were successfully [assembled] and [transformed] in E. coli, cultures were incubated overnight with () arabinose to induce reflectin expression.
- [Buffers] were prepared, and their pH checked and readjusted if necessary on the day of purification.
- An [inclusion body prep] was performed with 50ml of overnight culture. Reflectin was [purified] from the resulting lysate using a his-trap column.
- This procedure was repeated using a culture of the same bacteria from a different flask.
Results
Photospectroscopy readings from the eluted solution indicated that we had
