This animation will demonstrate how DNA microarray experiments are performed.
DNA microarrays, sometimes called DNA chips, reveal the expression of thousands of genes
on a solid surface, such as a microscope slide.
In this example, we’ll use yeast as a model system to illustrate one use of microarrays.
One common use of microarrays is to determine which genes are activated and which are repressed
when two populations of cells are compared.
Every gene is measured simultaneously.
As an example, we’ll compare what happens to yeast genes when cells are grown in aerobic
versus anaerobic conditions.
The cells grow and adjust which genes need to be activated or repressed in order to survive.
Now it is time to isolate the mRNA from both populations of cells.
The cells are spun in a centrifuge.
Now that the cells have gathered in pellets, we remove the liquid but not the cells.
Next, it is time to extract the mRNA from the cells.
When we add the extraction buffer, the mRNA is released into the solution.
Next, we remove the RNA and place it in a fresh tube.
Now, let’s make the cDNA from the mRNA.
Here we see three of the many mRNA molecules from each tube of cells.
Each mRNA is converted into red or green colored cDNA.
When the colored cDNA is made, the mRNA degrades.
Then we combine the red and green cDNA, mixing both colors into a single tube.
At last, it’s time to look at the DNA microarray.
In our experiment, a microarray or DNA chip contains about six thousand spots, each spot
is a different yeast coding sequence from a different gene.
Let’s choose three spots at random to follow in detail.
Each spot is made of DNA that can base pair with its complementary cDNA.
Here are partial sequences from each of the three spots we are observing.
Now let’s incubate the mixed cDNA with the DNA chip.
For the sake of our example, we’ll zoom in and show that some of the labeled cDNA
have bound to DNA in the spots and formed base pairings.
Here we see green and red cDNA bound to this spot.
Only red cDNA is bound to this spot.
And only green cDNA bound to this other spot.
In a real experiment, you would not see any of this detail; you would only see the original
microarray.
Now we must wash off the unbound cDNA to see what has bound to the microarray.
Let’s detect the bound cDNA so it can be visualized.
We begin by placing the microscope slide containing the microarray inside a scanner.
We’ll examine the next phase of the process, keeping our focus on the three spots we’ve
been following.
First, a green laser scans the microarray.
The resulting image is stored on a computer for later analysis.
Now it’s time for the red laser.
This image is also stored on a computer for later analysis.
Now we move to the analysis phase.
After we eject and safely store the microscope slide, we retrieve the red and green images
from the computer and create a merged visualization.
In the merged image we see an aerobic gene labeled in green, an anaerobic gene labeled
in red, and a gene labeled in yellow that was expressed in both aerobic and anaerobic
conditions.
This is one example of how DNA microarrays are used.
In an actual experiment, quantitative analysis would be conducted on all six thousand genes.