IGEM:MIT/2006/Notebook/2007-8-27

Things to Do Today

1. Make graphs with error bars from the plate reader data- DONE

2. Organize a meeting today with the advisors to discuss current events- DONE

3. Reserve 68-574 for meeting- DONE

4. Prepare slides for the meeting today- DONE

5. Make LCs of J45995, J45996, I7100 for experiment tomorrow- DONE

6. Develop protocol for salicylic acid/GFP experiment tomorrow

J45996/I7100 3-Hour Pulse

1. Add the following in 2 2-L flasks:

• 275 mL LB + Antibiotic

• 27.5 uL grown I7100 or J45996 culture

2. Grow the cultures up for 20 mins at 220 RPM at 37C

3. Lower the RPM for the cultures to 110 RPM at 37C. The start of this step is Time Zero.

4. At Time Zero, add .5 mL .1 M salicylic acid to a 125-mL flask and add 25 mL culture. Remove 3 200 uL aliquots for each culture for every 3 hours for hour 6, 9, 12, and 15.

6. After x hours (for x=4 to x=12 where x is even), extract 25 mL culture from master culture, pour into 125-mL flask, and add 0.5 mL .1 M salicylic acid to it for all cultures (grow up at 110 RPM). Also remove 3 200 uL aliquots for each culture and run on the plate reader.

7. At hour x+3 for hour x, remove 3 200 uL aliquots of each culture from the 2-L flask (with no salicylic acid added) and the 125-mL flask (with salicylic acid added).