Tools
This is the reading of the position of the four base pairs - adenine (A), cytosine (C), guanine (G)
and thymine (T) - in DNA to identify the positions and functions of genes.
The DNA molecule is too large to be sequenced in a single step. Using ultrasound or other mechanical means, the DNA is cut randomly into smaller fragments that are easier to read. As initially there are many DNA molecules, a multitude of fragments of different size are obtained, some of them having common sequences. Each fragment is read, the information is stored in a computer and powerful algorithms are used to reconstruct the complete DNA sequence.
DNA sequencing is also used to check whether the synthesis of a DNA molecule was carried out properly.
This is the "custom" manufacturing of DNA molecules in any order of base pairs, regardless if they belong to
living organisms or are sequences with no equivalent in nature. The difficulty of the synthesis increases
with the size of the DNA. The first step is a chemical synthesis of short sequences of hundreds or
thousands of base pairs. Then, larger sequences of thousands of base pairs are assembled by specific
enzymes. The final assembly of the synthetic DNA is made by molecular biology techniques involving
bacteria. The rate of DNA synthesis is limited by this final step and significant research efforts
have been done to automate it.
The first step is chemical synthesis of short sequences of hundreds or thousands of base pairs. Then, larger sequences of thousands of base pairs are assembled by specific enzymes. The final assembly of the synthetic DNA is achieved using molecular biology techniques involving bacteria. The rate of DNA synthesis is limited by this final step and significant research effort has been devoted to automating it.
Producing large amounts of DNA fragments quickly and at low cost, building and handling proto-cells
makes microfluidics a valuable tool for synthetic biology. At micron scale liquids like water have a
different behavior than at the everyday macroscopic scale: as their inertia is low one can accurately
monitor their mixing and movement to handle DNA sequences or proto-cells. In addition, working with
small amounts of liquid reduces reagent consumption and makes handling cheaper.
Particularly useful is the "lab on a chip", a plate of glass or plastic with microchannels where fluids move and perform several complex operations (transport, mixing, heating, results reading). In other words, it is a device that "shrinks" the laboratory to a few cm2 chip and allows the integration of multiple operations of the DNA or proto-cell synthesis.
Once synthesized, the DNA must be "inserted" in a biological frame, a favorable environment for its
operation and translation of genes into proteins. By analogy with computers, the synthetic DNA is the
software and the chassis is the hardware.
Many natural chassis are used today. They are living organisms whose initial DNA has been extracted: the bacterium Escherichia coli, host of our intestines, yeast, which makes beer and bread, or the harmless Bacillus subtilis, abundant in the soil. To function successfully the inoculated DNA should not be too different from the original DNA of the chassis, which severely limits the versatility of these solutions.
That is why there are attempts to use artificial systems called minimal cells. They have only a minimum of chemicals that enable them to function as a biological system. Minimal cells are easier to control but their construction is a big challenge.
Mathematical modeling and computer science: are necessary to process the large amounts of data produced by
DNA sequencing (the human genome has 3 billion base pairs), model and simulate complex interactions
between components of living organisms and, design and predict the behavior of biological systems
before building them.
A feature of synthetic biology is the use of engineering methods: beginning by defining the specifications (system characteristics), then designing the system by assembling standardized components listed in computer databases. This is followed by modeling (checking the system as a whole), implementation (system building) and validation testing (checking that specifications are met). These five steps can be repeated several times and it may be necessary to redefine the specifications, modify existing components or create new ones. Design and simulation require significant computational resources.
Standardization is specific to synthetic biology. In electronics, a radio is manufactured by
assembling various functional units (tuning circuit, amplifiers, oscillators, modulators) with well
defined functions. Each of them is build from standardized components such as transistors, resistors
and capacitors. Standardization allows different manufacturers to produce compatible components.
In addition, a functional unit that has been tested and validated also becomes a standard component
and can be used thereafter. Similarly, in synthetic biology one can design standardized and modular
devices and biological systems: DNA sequences are basic components, protein pathways are functional
blocks and so on.
To avoid contamination of natural organisms with genetic material from synthetic biology organisms,
the latter are physically isolated in confinement structures. Nevertheless, there are two other
methods. Trophic confinement consists in making synthetic organisms dependant on nutrients that
only the laboratory can provide. Semantic confinement is the design of synthetic organisms whose
genetic code or carrier of genetic information is different from natural organisms, thereby
preventing interference. Trophic and semantic confinements are now fields of intense research.
With these powerful tools, synthetic biology will pave the way for unprecedented medical and industrial applications, the same way that nanotechnology has led to numerous applications in electronics and materials engineering.
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