Bjorn's Experimental Log
December 14, 2005
- The first step of my adaptation protocol requires me to reproducibly grow bacteria to a concentration of 5e7 - 2e8 cells/mL in 1 hour
- Today I am taking one of my frozen aliquots of bacterial cells (BL21), thawing it, diluting it down to 2 different concentrations, and growing it back up to hopefully find out all the details of what initial concentration of cells will give me the desired concentration at the end of 1 hour.
- Going to try the following initial volumes of my frozen aliquot: 200uL, 500uL
- X uL of my Klett 80 (4e8 cells/mL) E. coli in ~9.9 mL LB solution in 125 mL flask
- Grow in orbital water bath at 37C, 200 rpm for 1 hr (or until back to Klett 80)
- Put flask on ice
- Serial dilute to get a 1e6 dilution of the Klett 80 solution and create 4 plates: 1x200uL, 2x100uL, 1x50uL
- Grow overnight in warm room (37C)
- After 1 hr, the 200uL sample was only at Klett 3 and the 500uL sample was at Klett 16. I let the Klett 16 sample continue to grow for another hour at which it was at Klett 36. At the end of one hour I still proceeded to plate and grow the 500uL sample to practice my technique.
December 15, 2005
- Remove my plates from the warm room
- Count colonies and calculate the true concentration of my Klett 16 solution
NOTE: I realized when I came in this morning that I diluted down my Klett 16 solutions as though they were of Klett 80, thus I had very few colonies on my plates this morning. Live and learn ehh???
Today I'm going to repeat yesterday's protocol but with an initial bacteria aliquot volume of 1 mL and 1.5 mL since I was not able to get enough bacteria grown at yesterday's initial concentrations.
RESULTS: After 1 hr the 1 mL sample grew to Klett 18 and the 1.5 mL sample grew to Klett 35. I diluted and plated them to grow overnight in the 37C room so I can count them in the morning.
December 16, 2005
Counted the colonies I grew yesterday and confirmed the following bacterial concentrations:
- 1.5 mL starting aliquot had 1.08e8 cells/mL
- 1.0 mL starting aliquot had 7.5e7 cells/mL
These results now let us know how much of the initial cell aliquot to add to the LB solution so we have btw 5e7 - 2e8 cells/mL after 1 hour of growth.
December 17-19, 2005
Been working on my multi-agent systems simulator all weekend. Its pretty cool ;)