BIOTECH Project
BIOTECH Home
Gateway to the BIOTECH Project
Laboratory Activities
What the BIOTECH Project can do in the classroom
Biotechnology Resources
Favorite resources online and in print
On Tour with BIOTECH
Follow the BIOTECH Project as it travels across Arizona
BIOTECH Bulletin Board
The latest news and your questions and comments
About the BIOTECH Project
What is the BIOTECH Project?
Name:
Date:
Period:
STUDENT GUIDE: Bacterial Transformation with Mystery
DNA
Some questions to get you thinking about today’s lab:
What can we use DNA for?
How can DNA be put into bacteria?
Why would we want to put DNA into bacteria?
How can we tell DNA is in the bacteria once we put it there?
What is a plasmid? What is ampicillin?
Materials for each group (students should work in groups of 4):
| Tube of mystery plasmid DNA (tubes numbered 1 or 2) |
1 tube of LB broth
|
| One tube of E. coli bacteria (on ice) |
micropipette
|
| 1 LB agar plate |
micropipette tips
|
| 1 LB agar plate with ampicillin for DNA (2 black stripes) |
small transfer pipet
|
| Q-tips or innoculation loops |
large transfer pipet
|
Materials to share:
Water bath at 42°C
ice
trash containers/biohazard waste bag
UV lights
Protocol:
1. Pipette the DNA solution from your numbered DNA tube into your E. coli bacteria tube and label the tube 1 or 2
2. Put tubes on ice for 5 minutes. Why do you think we put the tubes on ice?
3. In the meantime, each group should get one LB agar plate and one LB agar + ampicillin plate. Write your initials and your section number on your plates. You will be plating bacteria with DNA on an LB agar plate and on an LB agar +ampicillin plate. Mark these two plates with the DNA number on your tube. Where is the best place to label your plates? What is the control you are conducting?
4. Put tubes directly from ice into 42°C water bath for 50 seconds. What do you think heating the tubes does?
5. Put DNA tubes directly from water bath onto ice for 2 minutes.
6. With a large transfer pipet, add 0.25 ml LB broth into your DNA tube ( bring solution to the first mark on the pipet.) Incubate at room temperature for 10 minutes. What is the LB broth for?
7. With the small transfer pipet, pipet 0.15 ml (about half) from your DNA tube onto your LB agar plate and 0.15 ml onto your LB agar + ampicillin plate.
8. Spread the solutions on the plates using a sterile Q-tip. Be careful not to stab the agar.
9. Put your plates in a 37°C incubator for 24 hours. Why 37°C?
Challenge for Day 2: What is ampicillin and why do you think we used it in some of the plates?
What do you expect to grow on each of the plates?
|
LB agar
|
LB agar + ampicillin |
Do you expect to see any difference in bacterial growth on the two plates? | |
|
bacteria + DNA |
What do you think would have grown on these plates if no DNA had been added to these bacteria?
|
LB agar
|
LB agar + ampicillin |
Do you expect to see any difference in bacterial growth on the two plates? | |
|
bacteria (without DNA) |
Day 2:
What do you see on your plates?
Now look at your plates with UV light. What do you see?
Fill in the table with your data and the class data-
what does each group (#1 and 2) see on each type of plate?
|
Mystery DNA (number) |
LB agar
|
LB agar + ampicillin |
Based on the phenotype,
what is the DNA? |
|
#1
|
|||
|
#2
|
What does each DNA type (1 and 2) allow the bacteria to do?
BIOTECH Home | Laboratory
Activities | Biotechnology Resources
On Tour with BIOTECH | BIOTECH
Bulletin Board
|
BIOTECH Project Last Modified March 8, 2002 Nadja Anderson, Ph.D. |