Imperial College London/Notebook/2 October 2009

Harvard-GFP

 * There is a difference in fluorescence between 28°C and 37°C after compensating for OD.
 * Need to adjust the gain so that fluorescence doesn't saturate
 * 23rd September
 * 30th September

OD calibration

 * Preliminary results show that cell number varies with OD. However, the right dilution and enough replicates to overcome the huge variation are required.
 * 17th of September (up)
 * 23rd of September (awaiting results)
 * 25th of September
 * 2nd October (awaiting results)

Secondary carbon sources

 * There appears to be a plateau in cell number after 6 hours with 0.05% Glucose. However, more data is required to determine if this is the switch point.
 * 8th September (up)
 * 10th September (up)
 * 1st October

GFP pH

 * 18th September (awaiting data analysis)

IPTG growth

 * IPTG has no effect on cell growth under the concentrations tested (100uM to 5mM)
 * This is despit the dual effects of possible toxicity and protein production stress by IPTG on Lac-RFP constructs


 * 16th September (up)

Protocol considerations

 * Cells were grown overnight in 10ml of M9 media
 * The cells were allowed to grow until a high OD in the late morning, then placed in cold room
 * OD of around 0.4 to 1.2 was made up, and subsequently diluted up to 10-4 to 10-6
 * 100ul of cells were inoculated into each plate
 * The OD of the overnight cells were measured again after 3 hours on ice to check if cell number changes during dilution process

Results


[[Media:OD-calib02-10.xls| Download raw data]]

Conclusions

 * After 3 hours on ice, the OD stays relatively constant
 * Shows that number of cells during OD measurement and during plating stay the same. Therefore, OD correlates with the number of cells that are plated.

Suggestions/Improvements
Waiting to be uploaded