SEED/2010/Day 1

Morning

• EHS training

Afternoon

• Bench Inventory and Lab Tour
• Know your environment!
• Learn to pipet and use of bench equipment

Colony Streaking

This exercise will teach sterile technique and the important skill of being able to isolate single colonies. For this exercise, there are three cell types with three different antibiotic resistances (ampicillin, kanamycin, and chloramphenicol), and with three different color phenotypes (red, green, and no color). Each group will receive three tubes each of a single cell type but with an unknown antibiotic resistance. The first objective is to use these as standards to match antibiotic resistances with colors:

• Take one each of an Amp, Kan, and Cm plate and split each into thirds
• Streak each of the cell types on all three antibiotic plates

Each group will also receive three tubes that contain an unknown combination of the three different types of cells. The objective is to determine what each tube contains and to isolate single colonies from tubes that have a mix.

• Streak each of the three tubes on different LB plates with no antibiotic
  • The goal is to get single colonies
• Take another Amp, Kan, and Cm plate and split each into thirds
• Streak each of the unknowns on all the antibiotics to determine the mixture's composition

Pipetting

• Do dilutions of food coloring

Instructor Preparation

• Make enough LB plates with appropriate antibiotics (amp, kan, cm, and none). For each group:
  • 3 LB plates
  • 2 plates each of LB Amp, LB Kan, LB Cm
• Different E. coli with GFP, mCherry, or neither (no color) plasmids all with different antibiotics
• Mix liquid cultures with different combinations of these 3 cell types
• Some have only one, some have two types mixed together, and one may have all three mixed together

Instructor Post-lab

• grow plates and put into fridge