Name:_______________________________
Honors Biology Gene Chip Lab 2011
DNA Chips: From Genes to Evolution
Using DNA Chips (Microarrays) to Study Genes Involved in Evolution
Period:______
What you need to know/you might already know about gene chips:
 DNA microarrays are a powerful technology that scientists use to measure the transcription of thousands
of genes at a time.
 Even genes that are not highly expressed (transcribed into protein) may play an important role in the cell.
The LACK of expression of a gene may also play an important role in the cell. These chips are used to
measure expression, but it is up to the scientist to understand what the results mean!
Gene Chip Research Scenario:
A researcher was interested in the effects of high levels of CO2 (a potential problem associated with global
warming) on soybeans. She wanted to know which genes were activated under those conditions. The following
experiment was performed.
Control: Soybeans are grown under normal conditions
Experimental Group: Soybeans are grown with
high CO2 levels
active genes in both plants make mRNA which is extracted
Complimentary DNA (cDNA) is made from the extracted mRNA. Fluorescent nucleotides are used to do this
so the cDNA will be a unique color. Normal cDNA is dyed blue, while high C02 is red.
A gene chip is then prepared for the experiment:
Lots of single-stranded
DNA fragments are
fixed onto a microarray
slide. Each fragment
represents a specific
gene.
G
A
T
T
A
C
A
The cDNA from both samples is mixed and washed over the array. If a complimentary strand of DNA is on
the microarray, the two will bind.
This gene is being
expressed more in the
plants grown under
normal conditions.
This gene is
being
expressed more
in the plants
exposed to
high CO2.
No expression of
this gene by either
group of plants.
This gene is being
expressed in both the
control and the
experimental plants.
A computer can read the microarray and determine the degree of expression. The brighter the red spot, the more
it is being expressed in the experimental group relative to the same gene in the control group. The brighter the
blue spot, the more it is being expressed in the control group relative to the same gene in the experimental group.
Purple spots are being expressed equally by both groups of plants.
Created K. Gabric, 2005
**A colored version of this picture is available on the class website and in class, please reference it as you
are completing this pre-lab**
Use the link below to answer the pre-lab questions, you may have to read through the pre-lab and watch the video a
few times to ensure your understanding:
http://www.bio.davidson.edu/courses/genomics/chip/chip.html
1.
What is the difference between the two tubes of yeast cells that are grown?
2. Why is the liquid removed from the tubes after spinning?
3. What is suspended in the buffer?
4. Each tube is assigned a different color of cDNA. What is cDNA?
5. What three genes are being examined in this array?
6. What does it mean “incubate” the gene chip with the cDNA?
7. How is the DNA visualized? (How do we finally see something?)
8. Why is the green spot green?
9. Why is the red spot red?
10. Why is the yellow spot yellow?
11. Click on the grid to see real data, then click on the link for the “Whole Yeast Genome”. Describe what this
actual gene chip looks like.
12. In our lab, we will study gene expression in ancient DNA samples from different hominid organisms . The
cDNA from known Homo sapien relatives will be colored red, the cDNA from known Neanderthal species will
be colored blue. Using this information complete the following chart:
Gene Description
Color of Spot
A gene that is expressed in Homo sapiens
A gene that is expressed in yeast
A gene that is expressed in Neanderthals
A gene that is found in an organism
assumed to be descendents of only Neanderthals
A gene found in an organism assumed to be offspring
of both Homo sapien and Neanderthal
Introduction to the Simulation Scenario:
In this lab, you will perform a simulation that is very similar to the one that scientists use when conducting a real
DNA microarray experiment. The difference is that in a real lab, the DNA would be loaded by a computer and many
more samples of DNA could be tested at a time.
For this simulation we will study six genes found in various species:
Gene 1: Gene assumed to be found only in Homo sapiens
Gene 2: Gene found in common ancestor
Gene 3: Gene assumed to be found only in Neanderthals
Gene 4: Gene of unknown species
Gene 5: Gene assumed to be found only in Cro-magnon
Gene 6: Gene of unknown species
Materials:
 6 dropper bottles representing the various genes
 1 dropper bottle of hybridization solution (SAFETY WARNING: DO NOT GET IN EYES, ON SKIN, OR
ON CLOTHING, WEAR EYE PROTECTION THROUGHOUT THIS LAB)
 blank slides for preparing arrays
 Hot water bath for maintaining temperature of genes
 Goggles!!!
Procedure:
Step 1: Prepare the microarray slide
First, you will prepare (“print”) your DNA microarray by spotting each of the six different gene sequences onto a
glass slide. In real DNA microarrays, scientists also print thousands of microscopic DNA spots on glass slides, one
spot for each gene in the genome. Your Gene Chip will be much larger than a regular DNA microarray, and you will
be able to view them without specialized equipment.
1. Do not touch the surface of your slide (handle only on the edges). Using the china pencil, write your group
name on the label end of the slide.
2. Bring your labeled slide to the water bath area.
3. Using the dropper bottles, carefully spot the appropriate gene solutions into each of the labeled areas of
your slide. Be sure to place the correct DNA sequence in the correct spot. Once the sports are hardened
and dry, your microarray has been successfully “printed” with the 6 genes.
Step 2: Hybridize your microarray with labeled cDNA’s from sample species.
mRNA was isolated from different species and its cDNA was labeled with a blue or red dye. Neanderthal DNA is
blue, while Cro-Magnon DNA is red. You will not be able to see these dyes until the end of the experiment when
you visualize your results because the dyes are very diffuse.
You have been given a bottle (to be shared between groups) containing a solution of labeled cDNA, you cannot see
the color because the cDNA is very dilute. When added to the printed microarray slide, the labeled cDNA's in the
solution will base pair with the complementary DNA for each gene spotted onto the microarray. Binding cDNA to
the spotted DNA will produce a visable color.
1. Carefully drop 2-3 drops of the “hybridization solution” from the dropper bottle onto each spot.
2. DO NOT ALLOW the dropper bottle to touch the DNA spots.
Step 3: Visualize your labeled microarray results
1. Once you see some color appear, place the side on a sheet of white paper. You might be able to dunk the
slide in water to slow the color development if it gets too dark, but check with your teacher first.
2. Record your results by drawing the results below.
1
3
5
2
1
4
6
Analyze your results:
1. Which genes were expressed in the unknown samples? How do you know?
2. Which genes were not expressed in the unknown samples? How do you know?
3. Were there any samples that did not show any gene expression? Explain why this might have happened.
Explain what type of gene this might be.
4. Were there any genes expressed in more than one sample? Explain why this might have happened. Explain
what type of gene this might be.
5. Describe another microarray experiment that Wells would like to carry out. Be sure to include how he would
set up his experiment.