Team:PKU Beijing/Notebook/Protocol/Chemical Inducible GFP

Notebook > Protocol > Chemical Inducible Expression of GFP

Protocol for chemical inducible expression of GFP
 [[Media:PKU_Protocol_for_chemical_inducible_expression_of_GFP.pdf|download PDF version]]

Materials:
 * 4 groups of induce solution with a concentration gradient of 10^-7, 10^-5, 10^-3, 10^-2;
 * Overnight bacterial culture or bacterial colonies;
 * Phosphate Buffered Solution (PBS).

Procedure: 1. Add 20 μl of the overnight bacterial culture or pick a colony to 5ml of LB antibiotic medium, Incubate at 37 degree in a shaker till the OD600 value reaches 0.4-0.6. 2. Add 0.5 mL of the fresh bacterial culture and appropriate volume of inducer solution to prepare induction system with the concentration gradient of 10^-9, 10^-8, 10^-7, 10^-6, 10^-5, 10^-4, 10^-3, 10^-2. 3. Place the induction system at 37 degree for 2 hours. 4. Pellet bacterial cells by 4 min centrifugation at 4000 rpm, discard the supernatant. 5. Resuspend the pelleted cells in 500 μl of PBS. 6. Transfer 100 uL of bacterial resuspention into each well of 96-well plate to test the expression of GFP by flow cytometry or Microplate Reader.

Note: If desired, time sequential expression of GFP can also be tested, through verifying the incubating time of induction system at 37 degree.