Gene expression

Measurement

The expression of many genes is regulated after transcription (i.e., by agarose gel and hybridized to a radio-labeled RNA probe that is complementary to the target sequence. Northern blotting requires the use of radioactive reagents and can have lower data quality than more modern methods (due to the fact that quantification is done by measuring band strength in an image of a gel), but it is still often used. It does, for example, offer the benefit of allowing the discrimination of alternately spliced transcripts.

A more modern low-throughput approach for measuring mRNA abundance is real-time polymerase chain reaction or RT-PCR. With a carefully constructed standard curve RT-PCR can produce an absolute measurement such as number of copies of mRNA per nanolitre of homogenized tissue. The lower level of noise in data obtained via RT-PCR often makes this the method of choice, but the price of the required equipment and reagents can be prohibitive.

In addition to low-throughput methods, transcript levels for many genes at once (expression profiling) can be measured with concentration of different messenger RNAs. Recent advances in microarray technology allow for the quantification, on a single array, of transcript levels for every known gene in the human genome. The great advantage of tag-base methods is the "open architecture", allowing for the exact measurement of any transcript, known or unknown. Especially SuperSAGE recommends itself therefore also for studying organisms with unknown genomes.

Protein levels themselves can be estimated by a number of means. The most commonly used method is to perform a expression vector). Fusing a target protein to a reporter can also change the protein's behavior, including its cellular localization and expression level.

The pattern of detection of a gene or gene product may be described using terms such as facultative, constitutive, circadian, cyclic, housekeeping, or inducible.^[1]

Regulation of gene expression

Main article: Regulation of gene expression

Regulation of gene expression is the cellular control of the amount and timing of appearance of the functional product of a gene. Any step of gene expression may be modulated, from the DNA-RNA post-translational modification of a protein. Gene regulation gives the cell control over structure and function, and is the basis for cellular differentiation, morphogenesis and the versatility and adaptability of any organism.

Expression system

An expression system consists, minimally, of a source of expression vector, its cloned DNA, and the host for the vector that provide a context to allow foreign gene function in a host cell, that is, produce proteins at a high level".^[2][3]

In addition to these biological tools, certain naturally observed configurations of DNA (genes, promoters, enhancers, repressors) and the associated machinery itself are referred to as an expression system, as in the simple repressor 'switch' expression system in Lambda phage. It is these natural expression systems that inspire artificial expression systems, (such as the Tet-on and Tet-off expression systems).

Each expression system has distinct advantages and liabilities, and may be named after the host, the DNA source or the delivery mechanism for the genetic material. For example, common expression systems include bacteria (such as E.coli), yeast (such as S.cerevisiae), plasmid, artificial chromosomes, phage (such as lambda), cell lines, or virus (such as baculovirus, retrovirus, adenovirus).

Overexpression

In the laboratory, the protein encoded by a gene is sometimes expressed in increased quantity. This can come about by increasing the number of copies of the gene or increasing the binding strength of the promoter region.

Often, the DNA sequence for a protein of interest will be cloned or nuclear magnetic resonance experiments for structure determination.

Gene networks and expression

Main article: Gene regulatory network

Genes have sometimes been regarded as nodes in a network, with inputs being proteins such as transcription factors, and outputs being the level of gene expression. The node itself performs a function, and the operation of these functions have been interpreted as performing a kind of information processing within cell and determine cellular behaviour.

Techniques

• Primer: Used to facilitate expression
• Shuttle Vector

See also

• Bookmarking
• Expression profiling
• Expressed sequence tag
• Paramutation
• Sequence profiling tool
• Genetically modified organism
• Genetic engineering

References

1. ^ Glossary of gene expression terms
2. ^ cancerweb definition of expression system
3. ^ biology-online.org definition of expression system