Conclude growth curve

Continue with 1:10 000 dilutions until the concentration stabilizes. This occurs when two consecutive data points are approximately within ±25%.

Do a Clean program after each measurement.

Start a 30-minute countdown to remind you to collect the next sample.

1

Prepare the sample for BactoBox® measurement

1. Use a serological pipette to transfer a sample from the shake flask to a vial.

2. Use a permanent marker to label the vial cap with the sampling time.

3. Return the shake flask to the incubator shaker.

4. Start a 30-minute countdown to remind you to collect the next sample.

5. Vortex 30 seconds at max speed to disaggregate bacteria.

6. Prepare a 1:10 000 dilution using two sequential 1:100 dilution steps.

   1. First make a 1:100 dilution:

      1. Transfer 101 µL of your sample to 10 mL of diluent.

      2. Vortex 10 seconds at maximum speed.

   2. Then make the 1:10 000 dilution:

      1. Transfer 101 µL of the 1:100 dilution to 10 mL diluent.

      2. Vortex 10 seconds at maximum speed.

7. Transfer the tubing kit to the 1:10 000 vial.

2

Measure the diluted sample on BactoBox®

1. Click the Measure button in Access.

2. Add metadata:

   1. Dilution: 10 000.

   2. Label: E_coli_growth.

3

Clean the device

1. Transfer the tubing kit to the disinfection vial.

2. Click Clean.

Summary

You have now completed your growth curve experiment!

With these data you can calculate growth rate (µ), lag phase duration (ƛ), and carrying capacity (κ). These metrics will help you design of efficient seed trains.

Contact SBT if you need assistance on how to extract information from the growth curve experiment.