microarray

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