Microarray is a scientific method of
comparing the
expressed genes of
two different samples currently employed in
molecular biology.
What do you mean by "expressed genes"?
The easiest way to
understand this
concept is by following the thinking process outlined below.
1) It is a
fact that we all developed from a
fertilised egg.
2) That egg contains your
DNA, and from this one egg your
head, your
eyes, your
nose, your
fingers and your
toes developed.
3) Because we all developed from one fertilised
egg, therefore all the DNA in
every cell from your head to your toes contain
exactly the same DNA as that one egg carried.
4) Now
think: how come your NOSE does not appear on your TOES, but instead is found on your face? How come your heart
(containing
exactly the same DNA) is not on your
arm? And why don't you grow nails on your head instead of hair?
A
light bulb should be visible above your head about
now. You should have started to
understand what it means by the term
expressed genes. Carry on reading.
Just because all the cells in your body contain
exactly the same DNA
does not mean that all your cells have to
look the
same. The DNA comes with little
switches, which tell your cells what they are supposed to develop into and where they are to go. The swiches also turn the genes
on or
off, and regulate how
active they are (like the "
speed" button on a fan).
Genes are like little
light bulbs in the whole strand of DNA (which can be compared to a string of light bulbs you hang on your
Christmas tree). Genes can be turned on or off during
aging, in
different situations (like
stress or
illness), or in
different parts of the body.
If the DNA doesn't make the cells different, what does?
It is important to know what exactly goes on in the cell. Suppose we have a
gene for
antennae and that when we reach 11 years it will be turned on.
Antennae
------|----------|--------- .----.
off | 8 |
\__;_/
Age 10: turned off. We looked like this
Antennae \ /
------|----------|--------- .--V-.
ON | 8 |
\__;_/
Age 11: turned ON. We now look like this
Antennae
------|----------|---------
ON
E --- >
An enzyme E will come in to read the gene and copy it.
Antennae
------|----------|---------
ON
~~~~ E -- >
The enzyme makes a copy (the ~~~ thing) of the gene. This copy is known as mRNA.
Only genes that are copied (the scientific term is "transcribed") have
corresponding copies of
mRNA (messenger RNA) present in the cells. So how do you know which genes are active in the cell? Look at the
mRNA!
They're the ones that make the difference between your finger cells and your nose cells.
However, mRNA is
notoriously
unstable and degrades quickly. Therefore, scientists change mRNA into
cDNA in order to get a stable sample to work with.
cDNA is a copy of mRNA, except that it is in DNA format, not RNA format.
In summary, imagine a copier machine. You have a colourful book (
DNA) which you want to make copies of, but you don't want to copy all the pages. Only some pages are copied in colour. You may make 3 copies of page 5, 8 copies of page 9 and skip page 10. These
colour copies are your
mRNA. However, knowing the colour doesn't last, you decided to use your colour copies to make black-and-white copies as well. These black-and-white copies are your
cDNA.
How do you compare the cDNA of the two samples?
As I've said above, the
microarray compares the expressed genes (mRNA) of two different samples.
We allow the two samples to
mix on a glass slide, on which an
array (a whole lot of stuff that attracts the different genes) are printed on in astonishingly small dots. (Micro-array... get it?) A small
section of the slide looks like this.
-------------
| o o o o o |
| o o o o o |
| o o o o o |
| o o o o o |
| o o o o o |
-------------
One dot attracts only one specific gene.
The individual dots are full of small pieces of
oligos (singular: oligo), which act as
magnets attracting
cDNA to them.
-,-,-,-,-,- cDNA for antenna gene will bind to oligo
|
|
V
-'-'-'-'-'-'- oligo for the antenna gene (on glass)
----------------------- slide surface (glass)
The two samples are labelled with two
fluorescent dyes, one dye for each sample. The first dye is
red (think bright
luminescent red) and the second is
green (again, bright green).
Supposing we use the red dye to
label Sample R and the green to label sample G. We can then look at a single spot on the slide. If the spot is red in colour, it means that only Sample R
expresses that gene. If the spot is green, that means only sample G expresses that gene. And if the spot is
yellow, that means
both sample R and sample G express that gene in
equal amounts. Of course, there are other combinations of colour that enable you to know that both samples express that gene, but in
different amounts. An
empty spot would mean neither sample expressed that gene.
How do you perform a microarray?
A) Printing and processing the slides
Different oligos are
printed onto specially coated slides. The slides are then
processed so that the oligos
bind onto the surface of the slides, and any excess unbound oligos are washed off.
B) Preparation of the sample
RNA is first
extracted from the sample, and
converted to cDNA.
The cDNA is labelled with the appropriate
dye, and all excess dyes are washed away.
The labelled cDNA in
solution is then placed in contact with the
slide. This is known as hybridisation.
C) Scanning and evaluating the results
The slides are
scanned using a special slide scanner that can "
read" fluorescent dyes.
After the slides are scanned, the spots are
matched against a file that contain their true
identities. This is a lot of computer work. Each spot is identified as a certain gene.
The results are used to
generate a table, which tells us how much of each gene is expressed in one sample in
comparison with the other. The computer also does this; it generates the table by "viewing" the
relative amount of red dye compared to the amount of green dye it sees in one spot.
So what are microarrays good for?
Microarrays can be used in many fields of
biology. For example, a
scientist can compare the differences between a
virulent flu strain with a non-virulent flu strain and determine what makes that flu strain virulent. Drugs can also be used to
combat it.
In
developmental biology, a scientist can use microarrays to determine what genes swich on or off during the
growth
process.
In the
study of
diseases, it can be used to find out what genes the body turns on or off during the
onset of the illness and in order to overcome the illness.
Links and sources:
http://www.microarrays.org/
http://ihome.cuhk.edu.hk/~b400559/array.html
http://www.umich.edu/~retina/microarray.html -- Has a picture of a microarray