Preparation

kit of Promega (SVMinipreps)

incubative tube

1.5ml epp tube

MilliQ

Procedure

1. pour contents out of the incubative tube into the 1.5ml tube as you can
2. centrifuge for 10min (15,000rpm)
3. throw supernatant fluid away not to damage the precipitation

   ( you should decant by using yellow tip first / remove culture medium as you can throw waste water away in bio hazard!)

4. add 250μl cell resuspension solution (red label)、suspend completely

   (incomplete suspending decreases yields / you should use epp stand like a washboard)

5. add 250μl Cell lysis solution（green label）
6. turn the tube upside down four times slowly not to bubble
7. add 10μl Alkalin　Protease　Sol. （small bottle）
8. turn the tube upside down four times slowly not to bubble
9. wait for 5min （Be careful not to exceed 5min! colon bacillus will disintegrate too much!）
10. add 350μl Neutralization Sol. (blue label）
11. turn the tube upside down four times slowly not to bubble
12. centrifuge for 10min (15,000rpm)
13. put the supernatant fluid to column (germ’s wreckage is adhering below)
14. centrifuge for 1min (15,000rpm)
15. throw flow through (the liquid in the tube below) away
16. add 750μl Wash Sol. to column and centrifuge for 1min (15,000rpm)
17. throw flow through away, put 250μl Wash Sol. to column and centrifuge for 1min (15,000rpm)
18. change the column into 1.5ml tube and centrifuge for 2min (15,000rpm)
19. change the tube into new one and add 50μl MilliQ

    (use Nucleas-Free Water in the kit instead of MilliQ)

20. centrifuge for 1min (15,000rpm) after waiting for 1min
21. take 1 to 1.5μl and determine the concentration by NanoDrop (Don’t dilute)
22. label them