1. How to produce amylase by Bacillus Subtilis?

By submerge fermentation.
These enzymes have traditionally been obtained from submerged cultures because of ease
of handling and greater control of environmental factors such as temperature and pH
Bacillus subtilis was grown on nutrient agar at 37 C for 24 hours for inoculums preparation
The organism was propagated at 37C for 5 days in 25 ml of medium with shaking on a
shaker (150 rpm). Samples were taken at 12, 24, 48,72, 96 and 120 h. The supernatant of the
culture after centrifugation (10.000 rpm, 10 min) at 4C was used to determine amylase
activity.
The production of enzyme was directly related to the growth of the strain.

2. What is the media formulation for bacillus subtilis broth? Propose media preparation by
using peptone, magnesium sulphate pentahydrate, potassium chloride and starch