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