艾德華的部落格

質體純化步驟：

1. Incubate 50μl DH5α(with target gene) + 5ml LB medium(Km) @37℃ 150 r.p.m. 12hrs (or 14-16 hrs).
2. Centrifuge 1 ml Broth @ 13000r.p.m. 2min, then remove the supernatant.
3. Add 200 μl FAPD1 and disperse the pellet by pipetting.
4. Add 200 μl FAPD2 and rotate the eppendorf 10 times gently , then stand it for 2 min (until solution become transparent but don’t over 5 min).
5. Add 300 μl FAPD3 and rotate the eppendorf 10 times gently after that  centrifuge it @13000r.p.m. 8min.
6. Transfer the supernatant to a new eppendorf ,then centrifuge it @13000r.p.m. 8min.
7. Transfer the supernatant to a new combined column ,then centrifuge it @13000r.p.m. 30sec after that discard the liquid in the bottom.
8. Add 400 μl W1 buffer then centrifuge it @13000 r.p.m., 30 sec after that discard the liquid in the bottum.
9. Repeat step 8.
10. Add 600 μl Wash buffer then centrifuge it @13000 r.p.m., 30 sec after that discard the liquid in the bottum.
11. Centrifuge it again @ 13000 r.p.m., 3 min, then combine the upper column with a 1.5 ml eppendorf.
12. Stand it at 37℃ in the incubator for 10 min.
13. Add 50 μl hot water and stand for 2 min after that centrifuge it @13000 r.p.m. 2 min.
14. Collect the liquid in the bottom and add to the upper column again, then centrifuge it @13000 r.p.m. 2 min.
15. Finally, analyze the concentration、O.D.₂₆₀ O.D.₂₃₀ O.D.₂₈₀ by nanodrop。