BL21-AI

BL21 AI Expresion of toxic human proteins

1. Transform BL21 AI (Invitrogen) with pRARE2 plasmid. Plate on LB/Chl plate.

2. Transform BL21 AI/pRare2 with expression plasmid .

3. Grow in 1 L of TB media (12 g tryptone, 24 g yeast extract, 2.31 g KH2 PO4 (mono-basic), 12.5 g K 2 HPO4 (dibasic), 40 mL glycerol) containing
35 μg∕mL chloramphenicol and 50 μg∕mL kanamycin or 100  μg∕mL Amp at 37 °C until OD600 1⁄4 1.0.

4. Cool culture to 20 °C.

5. Add 4 mL of 50% w∕v arabinose and 1 M isopropyl-β-D-thiogalactopyranose (IPTG) to a final concentration of 0.4 mM.

6. Continue to culture for 1 hr at 20 °C.

7. Incubate the culture o/n at 20 oC.

8. Harvest cells by centrifugation.