Sonication

1. Take culture that has been grown up to 0.75 OD and centrifuge in the large centrifuge machine for 10 min at 4000rpm and 4*C.
2. Decant the supernatant and resuspend the pellet in 4ml of DI water.
3. Take the tube to the sonicator and sonicate for 30sec, then put on ice for 1 min and repeat. (make sure to sonicate with ethanol before each sample to avoid contamination!)
4. Centrifuge for an additional 10min at 4000rpm and 4*C.
5. Transfer the supernatant to a clean ON tube and discard the pellet

Assay

1. Add phytase (the supernatant) and 3mM of phytate in a 96 well plate in serial dilutions.
2. Use phytate only as a control.
3. Wait 20-30 minutes to allow the phytase to work on the phytate.
4. Add the reagent from the phosphate kit.
5. Wait 30 minutes to allow the reagent to work on the free phosphate.
6. Place the well plate into the plate reader and read at 650nm.