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| Gene Expression |
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Gene expression is a highly specific process in which a gene
is switched on, and therefore begins production of its protein.
Each cell in the human body (except for mature red blood cells and platelets, which do not contain a nucleus) contains a complete copy of the human genome: approximately 3.2 billion base pairs, organized onto 23 pairs of chromosomes (unpaired in gametes), with an estimated 50,000 genes.
Although every cell contains the full set of 50,000 genes, only
about one tenth of them are expressed, or turned "on," in any
particular type of cell. Cells look and act the way they do because
of the specific genes that they express and the amounts of gene products
produced. For example, a muscle cell, a skin cell, and a nerve
cell, could be distinguished by their gene expression profiles.
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Gene expression also varies within a certain type of cell at different
points in time. For example, prostate cells might express different genes at varying quantities in normal, precancerous, and malignant states:
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Two examples of technologies that facilitate the measurement of gene expression are:
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- Expressed Sequence Tags (ESTs)
Expressed Sequence Tags (ESTs) are short (usually about 300-500 bp),
single-pass sequence reads from the 5' and 3' ends of mRNA.
Typically ESTs are produced in large batches. They
represent a snapshot of genes expressed in a given
tissue and/or at a given developmental stage.
If a gene is very active, it will produce a large number of mRNAs. Therefore, the number of mRNA fragments from that gene in the library will be high compared to a less active gene.
In this way, ESTs are tags (some coding, others not) of mRNA, representing the genes that are being expressed in a given cell.
Note: mRNA -> cDNA:
mRNA is actually not a stable molecule for sequencing, so an enzyme called reverse transcriptase is used to convert mRNA back into a synthetic molecule called "cDNA," or complementary DNA.
cDNA libraries are built from different types of tissues, e.g., normal prostate, precancerous prostate, malignant prostate. The library contains short fragments of a sampling of the mRNAs produced in that tissue, and therefore represents the genes that are being actively transcribed.
The cDNA is sequenced and deposited into GenBank. However, the molecule type at the top of the GenBank record is still shown as mRNA, since that is the molecule that occurs in nature.
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- Micorarrays
"Microarray technology is one of several developing approaches
to comparatively analyze genome-wide patterns of mRNA expression. For mRNA expression studies, the ultimate goal is to develop arrays which contain
every gene in a genome against which mRNA expression levels can be quantitatively assessed." (From the NHGRI page, "About the Microarray Project")
Microarrays contain a wide sampling, or array, of cDNA from human genes. Then, DNA "probes" from a cell or tissue of interest are tagged with a flourescent label, and allowed to bind to the sample of human genes on the slide. Some of the cDNAs on the slide might not bind with any probes. That will be true if the gene represented by that cDNA is not being expressed by the cell of interest. On the other hand, some of the cDNAs will bind with a large quantity of flourescent probes, if that gene is highly expressed in the cell of interest. Other cDNAs will bind with a moderate quantity of probes.
The slides are put into a scanning microscope that can measure the brightness of each fluorescent dot; brightness reveals how much of a specific DNA fragment is present, an indicator of how active it is.
The NHGRI Glossary of Genetic Terms provides additional detail and an illustration. The NHGRI also is involved in a Microarray Project.
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