Soil samples collected from different sugarcane fields with
known history of extensive pesticide usage and located in Gurdaspur
district (Punjab, India) served as source of pesticide degrading
microbes. Representative soil samples (1 kg each) were collected
from four different fields were mixed thoroughly mixed. Pooled
soil sample (200 g) was suspended in 1 L of sterilized distilled
water with the help of a hand held blender and suspension filtered
by sieving through two layers of muslin cloth. The microbial biomass
in the filtrate was recovered in pellet by centrifugation at
10000 rpm for 10 min and resuspended in 0.5 mL of sterile distilled
water. For selective growth of only fipronil degrading microbes,
the mixed microbial suspension was grown in 50 mL of
Dorn’s broth containing fipronil (50 lgmL1) as sole source of carbon
and allowed to grow at 28 C on an orbital shaker (120 rpm).
After 96 h of growth, the procedure was repeated once by sub culturing
of 0.1 mL of the mixed culture into 50 mL of fresh Dorn’s
broth containing fipronil (50 lgmL1) and allowing it to grow as
above for 96 h to form enriched culture.