Template:SBB- Growth Rate Assay

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Growth Assay Protocol

To test how expression of parts affects the growth rate of cells, we want to compare growth of diluted cultures of arabinose-induced and non-induced cells.

For comparison between experiments, the control should be the strain without a plasmid, and the sample growth rates should be divided by the control growth rate to calculate a normalized growth rate. 0. Negative controls:

  • EC100D without plasmid (no AB resistance)
  • Bca1363 in EC100D

1. Grow overnight cultures of cells in appropriate antibiotic medium.

2. Dilute cells 1:10 for 500uL total volume in the blocks
2a. +/- arabinose, 100 µg/ml at the same time - immediately place into TECAN
2b. +/- arabinose, 100 µg/ml, let induce for 5 hours before proceeding

This should give starting OD of around 0.10, close to background of medium alone.

3. In a 96-well costar flat-bottomed black-sided plate, add 100 μL of DI water to every well on the outside edge. This will prevent evaporation of the experimental wells (in the middle of the plate).

4. Aliquot 100 of cells in triplicate into the remaining wells of the 96-well plate.

5. Run saved Tecan program "iGEM09 growth assay". Kinetic assay at 37C for 6hrs, reads at 20 min interval, shaking before each read.

6. When run is completed and before program is closed, go to Result details pulldown menu and save data so each well corresponds to a row with increasing time points. Copy and save data as excel file to import into MATLAB.