Bacterial Transformation Lab
Simulate a complete bacterial transformation experiment using the heat shock protocol. Adjust DNA concentration, heat shock parameters, and recovery time, then observe colony growth on selective agar plates and calculate transformation efficiency.
Guided Experiment: Effect of DNA Concentration on Transformation
How will changing the amount of DNA affect the number of transformed colonies? Will the relationship be linear or saturating?
Write your hypothesis in the Lab Report panel, then click Next.
Ready
Set parameters and run the experiment
Controls
Results
Data Table
(0 rows)| # | Trial | DNA(ng) | Heat Shock(s) | Recovery(min) | Colonies | Efficiency(/μg) |
|---|
Reference Guide
Heat Shock Protocol
The heat shock method uses temperature changes to create transient pores in the bacterial cell membrane, allowing DNA to enter the cell.
- Incubate bacteria + DNA on ice (CaCl₂ makes membranes competent)
- Heat shock at 42°C for 30-60 seconds (creates pores)
- Return to ice for 2 minutes (seals pores, trapping DNA inside)
- Add SOC media and recover at 37°C for 30-60 minutes
- Plate on selective media with antibiotic
Transformation Efficiency
Transformation efficiency measures how many bacteria successfully take up the plasmid DNA. It is expressed as colony-forming units per microgram of DNA.
Chemical transformation typically yields 10⁴ to 10⁶ transformants per μg DNA. Electroporation is more efficient, reaching 10⁹ to 10¹⁰ per μg.
Selectable Markers
Antibiotic resistance genes on the plasmid serve as selectable markers. When bacteria are plated on media containing the antibiotic, only cells that successfully took up the plasmid survive.
Without antibiotic selection (control plate), all viable bacteria grow, forming a lawn of colonies. This makes it impossible to distinguish transformed from non-transformed cells.
Common markers include ampicillin resistance (ampR, encoding beta-lactamase) and kanamycin resistance (kanR).
Experimental Controls
A well-designed transformation experiment includes several controls.
- Positive control — known plasmid DNA to verify the protocol works
- Negative control (no DNA) — competent cells plated without DNA to check for contamination
- No-antibiotic plate — to verify cell viability regardless of transformation
Variables you can optimize include DNA concentration, heat shock duration and temperature, recovery time, and competent cell preparation method.