Constructing Multispecies Biofilms from Rhizosphere Samples for Developing Advanced Biofertilizers

To begin, obtain cucumber seeds and disinfect them with 75%ethanol, followed by 2%sodium hypochlorite solution. After rinsing the seeds in sterile water, germinate them under sterile conditions. Transplant the consistently sprouted, two-leaf stage seedlings into pots containing two kilograms of black soil.

When the seedlings reach the four-leaf stage, invert the pot and isolate the roots along with two millimeter thick rhizosphere soil using sterilized gloves. Place the roots in 30 milliliters of PBSS buffer in a 50 milliliter centrifuge tube. Shake the tube for 20 minutes and filter the suspension through a 100 micrometer nylon cell filter to remove roots or large sediments.

Transfer the filtrate to another 50 milliliter centrifuge tube and centrifuge it at 3, 200 G for 15 minutes at room temperature. Temporarily store the tube at four degrees Celsius before extraction of rhizosphere soil bacterial cells.