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DNA Microarrays

If a microbiologist is studying bacteria that bioremediate, or break down, toxic wastes and wants to know which specific genes are active when that bacterium is degrading, say, PCBs, he would likely use a tool called the DNA microarray.

Microarrays enable scientists to monitor the activities of hundreds or thousands of genes at once. All microarrays (also called DNA chips or gene chips) work on the basic principle that complementary nucleotide sequences in DNA (and RNA) match up like the two halves of a piece of Velcro coming together.

Pattern of gene activity on a microarray chip.

A microarray consists of an orderly arrangement of bits of genetic material in super-tiny spots laid down in a grid on a suitable surface, often a glass slide with a specially chemically treated surface.

Each spot represents a single gene and contains millions of copies of that gene’s sequence made via PCR. A bacterium’s entire genetic make-up can be contained on a single gene chip. A computer keeps track of which gene is contained in each spot. There can be thousands or tens of thousands of these tiny spots on a single slide.

A specialized robotic machine uses super-thin stainless steel needles to dot the slide with the spots. The robot places the spots at precise intervals as it moves over the surface. The spots are incredibly miniscule, measured in micrometers (millionths of a meter); they typically range from 20 to 100 microns in diameter. Gene chips can be used for a number of purposes, but one of the most common is to determine which genes are expressed or activated under given conditions.

For example, a scientist might wish to figure out which genes in Streptococcus pneumoniae are involved in resistance to an antibiotic. He would first make or purchase a gene chip containing the genes for that bacterium. He would then break open and fish out the RNA from S. pneumoniae grown in plain media and from S. pneumoniae grown in media that contains a low level of the antibiotic (enough to encourage the bacteria to activate resistance genes but not enough to kill all the bacteria).

He would label, then tag the RNA from each sample with a different fluorescent dye. The plain media bacteria could be tagged with a dye that glows green, for example, and the RNA from the antibiotic-tainted media with a dye that glows red. The dye-tagged RNA bits from both bacteria groups would then be washed over the gene chip and left for a period of time to allow complementary strands to link up.

As the bits of RNA randomly bump into the DNA fixed to the chip, RNA sequences that are the complements to fixed DNA sequences latch onto the fixed material. Any unstuck, leftover bits of RNA are then washed off. The scientist uses a laser scanner to detect the fluorescent dyes and create a visual image of the pattern of the dyes.

Some of the gene spots might show up bright green under the scanner, indicating that those genes are active predominantly in bacteria without antibiotic resistance. Others glow yellow, indicating a mix of RNA from both the antibiotic-treated and untreated bacteria latched onto the DNA in those gene spots. This means those genes are active in both antibiotic-resistant and normal bacteria.

Some spots will glow red, however, and these spots indicate genes that are predominantly or only active in antibiotic-resistant bacteria. The brightness of any spot indicates how active those genes are. It’s the genes in the bright red spots that the scientist will now focus on in his efforts to thwart antibiotic resistance.

Because gene chips allow scientists to examine so many genes at once, they have greatly reduced the amount of time it takes to do experiments. Studies that once took months or even years to perform can now be done in a matter of days or even a few hours.



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