1346-A WSN 33 M15 88E
9/26/2012
Host Nico21(DE3)
induction: 0.3 mM IPTG, 6 hours, 37 oC
(expression is not good: 1 mM IPTG, o/n at 37 oC, or 0.3 mM IPTG, 30 oC o/n.
7 L culture.
Ni column, then gel filtration.
Gel filtration
Buffer 50 mM Tris-HCl, pH8.0, 200 mM NaCl.
column:
Flow rate: 1.5 to 5 ml., 4ml/tube
Run 10 ul on Bis-Tris gel
9/27/2012
Left gel:
| 1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
9 |
10 |
11 |
12 |
13 |
14 |
15 |
16 | |
| Fraction |
20 |
21 |
22 |
23 |
24 |
19 |
20 |
21 |
22 |
23 |
24 |
25 |
20 |
22 |
22 |
23 |
| OD280 |
0.3 |
0.68 |
0.80 |
0.50 |
0.20 |
0.30 |
1.61 |
2.43 |
1.33 |
0.72 |
0.24 |
0.13 |
1.35 |
2.82 |
1.98 |
0.31 |
combine tube 20, 21,22,23-->OD280=1.6, total ~140 ml
combine other
Right gel:
| 1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
9 |
10 |
11 |
12 |
13 |
14 |
15 |
16 | |
| Fraction |
C5 |
C6 |
C7 |
C8 |
C9 |
C10 |
D10 |
D9 |
D8 |
D7 |
D6 |
D5 |
D4 |
D3 |
D2 |
D1 |
| OD280 |
0.63 |
0.68 | 0.69 |
0.72 |
0.77 |
0.84 |
0.97 |
1.0 |
1.1 |
1.2 |
1.6 |
2.0 |
1.8 |
1.5 |
1.2 |
1.0 |