Measuring growth rates

The first question we will try and answer is, how fast does E. coli grow under standard conditions?

We specify "standard conditions" as accurately as possible so that other people, working in other places, can repeat our observations.   Even minor (an unappreciated) differences can sometimes make a big difference.

For E. coli, standard conditions are LB media, at 37°C with constant rotation; the rotation insures that the culture is aerobic. 

Each 5 ml culture is started from a single colony, isolated from an agar plate.  As you might expect, the exact number of bacteria used to inoculate each culture varies from culture to culture.

Take care to use the vSPEC correctly.  Just like a physical spectrophotometer, it drifts over time and so must be recalibrated often.  Begin by

• selecting a time
• then check that the dark current is zero
• check the blank produces an A=0

If you do not do these things, your measurements will be inaccurate. Inaccuracy is bad!

The wavelength selector on your vSPEC has been set to 600 nm.  If you find it difficult to use the knobs, you can use the autoZero and autoBlank buttons.

Record your time and absorbencies readings using the vDatapad. Once your measurements are made, press the "plot data" button to produce a graph of your data.

Use the arrow cursors to select the region of the A versus time (T) curve - the doubling time will be determined automatically, through a least-squares curve fit of the region you chose.

HERE'S A HINT: If you include too many points where A is close to zero or greater than 0.6, your estimate of the doubling time will be inaccurate.

WRITE-UP

Remember your question!

Results: Once you succeed in accurately determining the doubling time of your culture, print out your data graph.  

Discuss: Assuming that all of the students in the class are actually using the same strain of E. coli and the same culture media:

• Q1: What would happen to your growth rate if you did not rotated the culture (and why)?
• Q2: Why are there differences between the growth rate curves you measure in your first and second cultures? 
• Q3: What is the source of variation in your studies?
• Q4: Two substrains show different doubling times under standard conditions; provide a testable hypothesis for why that might be.