What a Colorful World

Simplifying assumptions about “the cell” are brought into question when different strains are transformed with DNA that makes them grow in colorful ways.

In this lab, two closely related bacterial cell types are transformed with two color-generating plasmids. This content is relevant to genetics and inheritance lessons, molecular biology, and DNA expression units. Sterile technique and microbial culturing are emphasized in this streamlined and effective alternative to more common transformation protocols.The engineering concept of “chassis selection” gives this lab a context beyond a simple exploration of the transformation technique. Because the pigments that are generated by the plasmids are visible to the naked eye, it’s easy to discern if the expression differs between the two chassis, and the data can be analyzed both quantitatively and qualitatively. Students can design further experiments to understand what differences between chassis that might explain their data.

BACKGROUND

Relevant content and skills

Content

DNA Basics: what it is and how it works
Gene Expression: Transcription, Translation, Operons
Recombinant DNA: Restriction enzymes, cloning, selection with antibiotic resistance

Skills

Pipetting
Timing Reactions
Interpreting Data
Generating Hypotheses
Basic microbiology skills: Culturing bacteria, Aseptic technique, Selection through antibiotic resistance

SOME LAB MATERIALS TO HAVE ON HAND

HARDWARE: Some basic biology laboratory equipment such as glassware, pipets, a water bath or heat block to heat samples, an icebucket or styrofoam cup to cool samples, 4C fridge and ice-cold freezer (ideally to -20C) to store samples, a stir plate to mix samples, and an incubator to grow samples (though room temperature can work too). Some additional experiments might need a chemical hood, electrophoresis chambers, spectrophotometer, or a PCR machine.
CONSUMABLES: Some basic consumable laboratory equipment such as pipet tips, plastic tubes, growth media, petri dishes, toothpicks, spreaders, sterile loops, bleach or the like for decontamination and disposal of bioreagents. Some additional experiments might need additional consumables such as gel running buffers, loading buffers, stains to visualize DNA or proteins, cuvettes, PCR master mix, or an autoclave for sterilizing reagents.

RESOURCES AND DOWNLOADS

Specific materials for teaching this lab

CLASSROOM RESOURCES

READINGS

PPTs

VIDEOS

HANDOUTS

LAB RESOURCES

LAB MATERIALS

VIDEOS

PPTs

Lab Intro v1

OTHER RESOURCES

Workflow for the Colorful World lab

INTERPRETING THE RESULTS

A starting place for analysis and feedback

Download the teacher’s version of the lab manual for MUCH more information about the experimental outcomes, as well as pre- and post-lab questions to assess student understanding.

Briefly Summarized:

We generally observe strain 4-1 produces large, light green colonies and dark purple colonies, while strain 4-2 usually produces dark, small green colonies and no purple colonies.

We generally observe transformation efficiencies around 1*10^3 colonies/microgram of DNA. However, variations in the protocol, such as incubation at room temperature, may produce different results.

To achieve high transformation efficiency and distinct color differences, the students must be precise when conducting the transformation protocol. For instance, a water bath in excess of 42°C or cells left in the bath for more than 90 seconds may damage the cells and adversely affect the transformation efficiency.

As a control for this experiment, some teachers like to plate a “no DNA” control on an LB petri dishes (you’d expect a lawn of cells). Some like to plate the “+DNA” samples on LB also, in which case you’d see a lawn but ~none in that lawn will be colorful since the frequency of transformation is so low.