3 - Bacterial growth

Grow bacteria

Streak frozen cells on agar plate

Pick a colony and grow overnight in liquid LB (plus antibiotic if necessary)

Concentrate ON culture in centrifuge

Make 2 dilution series: 10-fold, 2-fold

• 2 fold

  • tubes 1A - 10A
  • 0.5 mL LB in each
  • add .5 mL culture to first, mix thoroughly
  • serially dilute by moving 0.5 mL along the series*

• 10 fold

  • tubes 1B - 10B
  • 0.9 mL LB in each
  • add .1 mL culture to first, mix thoroughly
  • serially dilute by moving 0.1 mL along the series

Take OD measurements

• Load 250 uL of each dilution into 96 well plate
• samples in 1st 10 wells of 1st 2 rows
• LB in last 2 wells
• Read, using bug OD

Plot results

• Put fraction of original culture next to each OD
• Cut, paste special, values only
• Sort data
• Make a scatterplot
• Find the lowest concentration just above the noise for hemocytometer

Hemocytometer

• Load 10 uL into hemocytometer.
• Photograph.
• Also take photographs of too concentrated and too dilute samples

Colony Counts

• spread 100 uL from last 5 10-fold dilutions on agar plates
• Grow overnight
• Photograph

Part 1 of PowerPoint from 2020 remote lab

Part 2 of PowerPoint from 2020 remote lab