1. By: Lynn More - Olympian High School and UCSD
Protein Transformation
Lab Preview
UCSD: BioBridge Program
By: Lynn More - Olympian High School and UCSD

2. Materials Checklist Students make bags in groups of four:
2 empty microcentrifuge tubes
4 disposable transfer pipette
1 Inoculating loop
6 cotton swabs
Teacher will have at lab tables for each group of four:
1 waterproof pen - for labeling
1 LB plate
2 LB/AMP plates
Ice bucket (cup with ice and water)
1 microcentrifuge tube of CaCl2 on ice
1 microcentrifuge tube of plasmid labeled either PM1 or PM2 on ice.
2 Lab station waste containers - one with 10% bleach, other empty
Tape for sealing plates after inoculation
Timer

3. 1. Untape petri dishes and put them on the placemat where they go.
Save the tape for the end!
2. Label equipment
3. Perform Transformation Lab
4. Incubate your plates
5. Turn in your group’s “What Do You Know” sheet

4. Labeling Equipment + and Table letter - and Table letter LB/Amp +
Period & Table
LB/Amp -
Period & Table
LB/ no Amp -
Period & Table

5. Equipment
0.5 mL
Transfer Pipette
0.25 mL
0.1 mL
Microcentrifuge Tube

6. FP Transformation procedure
Suspend bacterial colonies in CaCl2
Add plasmid DNA
Incubate on ice for 10 minutes
Work on you’re the “What Do You Know?” info

7. FP Transformation procedure
Suspend bacterial colonies in CaCl2
Add plasmid DNA
Heat-shock:
• ice for 10 minutes
• 42°C for 45 seconds
• back on ice again for 2 min
Plate out bacteria
Read & Follow your lab instructions!

8. Transformation Flow Chart+ -
CaCl2 +
Bacteria
Plasmid
(PM1 or PM2)
CaCl2 +
Bacteria
Incubate on ice for 10 minutes
Heat Shock: Ice – Heat - Ice
LB/Amp +
LB/Amp -
LB/No Amp -
Experimental Plate:
Colonies of bacteria
With plasmids
Control Plate: LB/Amp-:
Testing effectiveness
Of Ampicillin
And contamination
Control Plate: LB/No Amp-:
“Lawn” testing viability
of the bacteria
and the nutrient
Purpose of
Each plate

9. What is in the plate to start with? What do you expect to see?
What do you know?
Plate
What is in the plate to start with?
What do you expect to see?
Experimental Plate:
LB/Amp +
Control Plate:
LB/Amp -
Control Plate:
LB/No Amp -

10. What is in the plate to start with? What do you expect to see?
What do you know? (KEY)
Plate
What is in the plate to start with?
What do you expect to see?
• Luria Broth (Nutrient/Food)
• Ampicillin (antibiotic)
(tainted food)
• transformed bacteria
(bacteria + the plasmid)
Glowing bacteria survive
(regular bacteria die off)
transformed bacteria
Colored bacteria
• regular bacteria
(bacteria minus plasmid)
No growth at all
(ampicillin kills off the regualr bacteria like it’s supposed to)
• Regular food, Food is NOT tainted
A lot of growth (lawn)
(give regular bacteria good food and it’ll GROW!)
Experimental Plate:
LB/Amp +
Control Plate:
LB/Amp -
Control Plate:
LB/No Amp -