Tools and Technologies

Here is an overview of the major technologies and tools used in biotechnology

Bioprocessing Technology
The oldest of the biotechnologies, bioprocessing, uses living cells or the molecular components of cells’ manufacturing machinery to produce desired products. The living cells most commonly used are one-celled microorganisms, such as yeast and bacteria; the biomolecular components used include DNA (which encodes the cells’ genetic information) and enzymes (proteins that catalyze biochemical reactions).

A form of bioprocessing, microbial fermentation, has been used for thousands of years to brew beer, make wine, leaven bread and pickle foods. In the mid-1800s, when we discovered microorganisms and realized they were responsible for these useful products, we greatly expanded our use of microbial fermentation. We now rely on the remarkably diverse manufacturing capability of naturally occurring microorganisms to provide us with products such as antibiotics, birth control pills, vaccines, amino acids, vitamins, industrial solvents, pigments, pesticides, biodegradable plastics, laundry-detergent enzymes and food-processing aids.

Cell Culture
Cell-culture technology is the growing of cells outside of living organisms (ex vivo).

Plant Cell Culture
An essential step in creating transgenic crops, plant cell culture also provides us with an environmentally sound and economically feasible option for obtaining naturally occurring products with therapeutic value, such as the chemotherapeutic agent paclitaxel, a compound found in yew trees and marketed under the name Taxol®. Plant cell culture is also under study as a manufacturing tool for therapeutic proteins, and is an important source of compounds used as flavors, colors and aromas by the food-processing industry.

Insect Cell Culture
Insect cell culture can broaden our use of biological-control agents that kill insect pests without harming beneficial ones or having pesticides accumulate in the environment. Even though we have recognized the environmental advantages of biological control for decades, the manufacture of such products in marketable amounts has been impossible. Insect cell culture removes these manufacturing constraints.
Like plant cell culture, insect cell culture is being investigated as a production method of therapeutic proteins. Insect cell culture is also being investigated for the production of VLP (virus-like particle) vaccines against infectious diseases such as SARS and influenza, which could lower costs and eliminate the safety concerns associated with the traditional egg-based process. A patient-specific cancer vaccine, Provenge, that utilizes insect cell culture is up for FDA approval, along with a second vaccine for Human Papilloma Virus (HPV), Cervarix.

Mammalian Cell Culture
Livestock breeding has used mammalian cell culture for decades. Eggs and sperm, taken from genetically superior cows and bulls, are united in the lab, and the resulting embryos are grown in culture before being implanted. A similar form of mammalian cell culture has also been an essential component of the human in vitro fertilization process. Our use of mammalian cell culture now extends well beyond the brief maintenance of cells in culture for reproductive purposes. Mammalian cell culture can supplement—and may one day replace—animal testing of medicines. As with plant cell culture and insect cell culture, we are relying on mammalian cells to synthesize therapeutic compounds, in particular, certain mammalian proteins too complex to be manufactured by genetically modified microorganisms. For example, monoclonal antibodies are produced through mammalian cell culture. Scientists are also investigating the use of mammalian cell culture as a production technology for influenza vaccines. In 2006, the Department of Health and Human Services awarded contracts totaling approximately $1 billion to several vaccine manufacturers to develop new cell-culture technologies for manufacturing influenza vaccine. Cell-culture technology has been used for other vaccines, but each vaccine process is unique and influenza vaccine manufacturing has traditionally been performed using large quantities of eggs. New manufacturing technologies are an essential part of pandemic influenza preparedness and require extensive research and development. Cell-culture techniques could enhance the manufacturing capabilities and capacity.

Recombinant DNA Technology
Recombinant DNA is the foundation of modern biotechnology. The term recombinant DNA literally means the joining—or recombining— of two pieces of DNA from different sources, such as from two different organisms.

Humans began to change the genetic material of domesticated plants and animals thousands of years ago by selecting which individuals would reproduce. By breeding individuals with valuable genetic traits while excluding others from reproduction, we changed the genetic makeup of the plants and animals we domesticated. Now, in addition to using selective breeding, we recombine genes at the molecular level using the more precise techniques of recombinant DNA technology. Making manipulations more precise and outcomes more certain, biotechnology decreases the risk of producing organisms with unexpected traits and avoids the time-consuming, trial-and error approach of selective breeding. Genetic modification through selective breeding and recombinant DNA techniques resemble each other, but there are important differences:

●Genetic modification using recombinant DNA techniques allows us to move single genes whose functions we know from one organism to another.
● In selective breeding, large sets of genes of unknown function are transferred between related organisms.

Techniques for making selective breeding more predictable and precise have been evolving over the years. In the early 1900s, Hugo DeVries, Karl Correns and Eric Tshermark rediscovered Mendel’s laws of heredity. In 1953, James Watson and Francis Crick deduced DNA’s structure from experimental clues and model building. In 1972, Paul Berg and colleagues created the first recombinant DNA molecules, using restriction enzymes. Ten years later, the first recombinant DNA-based drug (recombinant human insulin) was introduced to the market. By 2000 the human genome had been sequenced and today we use recombinant DNA techniques, in conjunction with molecular cloning to:

●● produce new medicines and safer vaccines.
●● enhance biocontrol agents in agriculture.
●● increase agricultural yields and decrease production costs.
●● reduce allergy-producing characteristics of some foods.
●● improve food’s nutritional value.
●● develop biodegradable plastics and other biobased products.
●● decrease water and air pollution.
●● slow food spoilage.

Monoclonal Antibodies
Monoclonal antibody technology uses immune-system cells to make proteins called antibodies, which help the body to destroy foreign invaders such as viruses or bacteria. We have all experienced the extraordinary specificity of antibodies (specificity refers to the ability of antibodies to bind to only one type of molecule). For example, the antibodies that attack a flu virus one winter may do little to protect us from a slightly different
flu virus the next year.

The method of making monoclonal antibodies involves fusing a human myeloma cell (a cancerous immune B cell) that can no longer secrete antibodies to a normal B cell from a mouse that has been immunized to secrete a particular antibody. The myeloma component helps the hybrid cell multiply indefinitely, and the fused cell—called a hybridoma—can be cultured. The cells all produce exactly the same antibody—hence the term monoclonal antibody. As with the antibodies our bodies make to fight disease, monoclonal antibodies bind with specificity to their targets, making them tempting candidates for fighting cancer, infections and other diseases. The specificity of antibodies also makes them powerful diagnostic tools. They can locate substances that are present in minuscule amounts and measure them with great accuracy. For example, monoclonal antibodies can be used to:

●● locate environmental pollutants.
●● detect harmful miroorganisms in food.
●● distinguish cancer cells from normal cells.
●● diagnose infectious diseases in humans, animals and plants more quickly and more accurately than ever before.

In addition to their value as detection devices, monoclonal antibodies (MAbs) can provide us with highly specific therapeutic compounds. Monoclonal antibodies can treat cancer, for example, by binding to and disabling a crucial receptor or other protein associated with cancerous cells. Joined to a toxin, a monoclonal antibody can selectively deliver chemotherapy to a cancer cell while avoiding healthy cells. Monoclonal antibodies have also been developed to treat organ-transplant rejection and autoimmune diseases by specifically targeting the type of immune system cell responsible for these attacks.

Monoclonal antibodies can be created in mouse cells, but often the human patient mounts an immune response to mouse antibodies. This immune response not only eliminates the herapeutic MAb administered, but is also dangerous for patients and may cause lasting damage. To reduce this problem scientists create chimeric, or humanized, antibodies in which some parts of mouse origin are replaced with parts of human origin. Such antibodies are less likely to trigger an unwanted immune response.

Cloning
Cloning technology allows us to generate a population of genetically identical molecules, cells, plants or animals. Its applications are extraordinarily broad and extend into many research and product areas. Any legislative or regulatory action directed at “cloning” must take great care in defining the term precisely so that the intended activities and products are covered while others are not inadvertently captured.

Molecular or Gene Cloning
Molecular or gene cloning, the process of creating genetically identical DNA molecules, provides the foundation of the molecular biology revolution and is a fundamental tool of biotechnology. Virtually all applications in biotechnology, from drug discovery and development to the production of transgenic crops, depend on gene cloning. The research findings made possible through molecular cloning include identifying, localizing and characterizing genes; creating genetic maps and sequencing entire genomes; associating genes with traits and determining the molecular basis of these traits.


Animal Cloning
Animal cloning has been rapidly improving livestock herds for more than two decades and has been an important tool for scientific researchers since the 1950s. Although the 1997 debut
of Dolly the cloned sheep was a worldwide media event, animal cloning was not altogether new. Dolly was considered a scientific breakthrough not because she was a clone, but because the source of the genetic material used to produce Dolly was an adult cell, not an embryonic one. There are, in fact, two ways to make an exact genetic copy of an organism such as a sheep or a laboratory mouse:

●● Embryo Splitting is the old-fashioned way to clone. Embryo splitting mimics the natural process of creating identical twins, only in a Petri dish rather than the mother’s womb. Researchers manually separate a very early embryo into two parts and then allow each part to divide and develop on its own. The resulting embryos are placed into a surrogate mother, where they are carried to term and delivered. Since all the embryos come from the same zygote, they are genetically identical.
●● Somatic cell nuclear transfer (SCNT) starts with the isolation of a somatic (body) cell, which is any cell other than those used for reproduction (sperm and egg, known as the germ cells). In mammals, every somatic cell has two complete sets of chromosomes, whereas the germ cells have only one complete set. To make Dolly, scientists transferred the nucleus of a somatic
cell taken from an adult female sheep to an egg cell from which the nucleus had been removed. After some chemical manipulation, the egg cell, with the new nucleus, behaved like a freshly fertilized zygote. It developed into an embryo, which was implanted into a surrogate mother and carried to term.
Animal cloning provides many benefits. The technology can help farmers produce animals with superior characteristics, and it provides a tool for zoo researchers to save endangered species.
Also, in conjunction with recombinant DNA technologies, cloning can provide excellent animal models for studying genetic diseases and other conditions such as aging and cancer. In the future, these technologies will help us discover drugs and evaluate other forms of therapy, such as gene and cell therapy.

Protein Engineering
Protein engineering technology is used, often in conjunction with recombinant DNA techniques, to improve existing proteins (e.g., enzymes, antibodies and cell receptors) and create proteins not found in nature. These proteins may be used in drug development, food processing and industrial manufacturing. Protein engineering has most commonly been used to alter the catalytic properties of enzymes to develop ecologically sustainable industrial processes. Enzymes are environmentally superior to most other catalysts used in industrial manufacturing because, as biocatalysts, they dissolve in water and work best at neutral pH and comparatively low temperatures. In addition, because biocatalysts are more specific than chemical catalysts, they also produce fewer unwanted byproducts. Makers of chemicals, textiles, pharmaceuticals, pulp and paper, food and feed, and energy are all benefiting from cleaner, more energy-efficient production made possible with biocatalysts.

The characteristics that make biocatalysts environmentally advantageous may, however, limit their usefulness in certain industrial processes. For example, most enzymes fall apart at high temperatures. Scientists are circumventing these limitations by using protein engineering to increase enzyme stability under harsh manufacturing conditions. In addition to industrial applications, medical researchers have used protein engineering to design novel proteins that can bind to and deactivate viruses and tumor-causing genes; create especially effective vaccines; and study the membrane receptor proteins that are so often the targets of pharmaceutical compounds.

Food scientists are using protein engineering to improve the functionality of plant storage proteins and develop new proteins as gelling agents. In addition, researchers are developing new proteins to respond to chemical and biological attacks. For example, hydrolases detoxify a variety of nerve agents as well as commonly used pesticides. Enzymes are safe to produce, store and use, making them an effective and sustainable approach to toxic materials decontamination.

Biosensors
Biosensor technology couples our knowledge of biology with advances in microelectronics. A biosensor is composed of a biological component, such as a cell, enzyme or antibody, linked to a tiny transducer—a device powered by one system that then supplies power (usually in another form) to a second system. Biosensors are detecting devices that rely on the specificity of cells and molecules to identify and measure substances at extremely low concentrations. When the substance of interest binds with the biological component, the transducer produces an electrical or optical signal proportional to the concentration of the substance. Biosensors can, for example: measure the
●●nutritional value, freshness and safety of food.
●● provide emergency room physicians with bedside measures of vital blood components.
●● locate and measure environmental pollutants.
●● detect and quantify explosives, toxins and biowarfare agents.

Nanobiotechnology
Nanotechnology is the next stop in the miniaturization path that gave us microelectronics, microchips and microcircuits. The word nanotechnology derives from nanometer, which is one-thousandth of a micrometer (micron), or the approximate size of a single molecule. Nanotechnology—the study, manipulation and manufacture of ultra-small structures and machines made of as few as one molecule—was made possible by the development of microscopic tools for imaging and manipulating single molecules and measuring the electromagnetic forces between them.
Nanobiotechnology joins the breakthroughs in nanotechnology to those in molecular biology. Molecular biologists help nanotechnologists understand and access the nanostructures and nanomachines designed by 4 billion years of evolutionary engineering—cell machinery and biological molecules. Exploiting the extraordinary properties of biological molecules and cell
processes, nanotechnologists can accomplish many goals that are difficult or impossible to achieve by other means. For example, rather than build silicon scaffolding for nanostructures,
DNA’s ladder structure provides nanotechnologists with a natural framework for assembling nanostructures. That’s because DNA is a nanostructure; its highly specific bonding properties
bring atoms together in a predictable pattern on a nano scale.

Nanotechnologists also rely on the self-assembling properties of biological molecules to create nanostructures, such as lipids that spontaneously form liquid crystals. Most appropriately, DNA, the information storage molecule, may serve as the basis of the next generation of computers.
DNA has been used not only to build nanostructures but also as an essential component of nanomachines.
Most appropriately, DNA—the information storage molecule—may serve as the basis of the next generation of computers. As microprocessors and microcircuits shrink to nanoprocessors and nanocircuits, DNA molecules mounted onto silicon chips may replace microchips with electron flow-channels etched in silicon. Such biochips are DNA-based processors that use DNA’s extraordinary information storage capacity. (Conceptually, they are very different from the DNA microarray chips discussed below.) Biochips exploit the properties of DNA to solve computational problems; in essence, they use DNA to do math. Scientists have shown that 1,000 DNA molecules can solve in four months computational problems that would require a century for a computer to solve. Other biological molecules are assisting in our continual quest to store and transmit more information in smaller places. For example, some researchers are using light-absorbing molecules, such as those found in our retinas, to increase the storage capacity of CDs a thousand-fold.

Some applications of nanobiotechnology include: increasing the speed and power of disease diagnostics.
●● creating bio-nanostructures for getting functional molecules into cells.
●● improving the specificity and timing of drug delivery.
●●miniaturizing biosensors by integrating the biological and
electronic components into a single, minute component.
●● encouraging the development of green manufacturing practices.

Microarrays
Microarray technology is transforming laboratory research because it allows us to analyze tens of thousands of data points simultaneously. Thousands of DNA or protein molecules, or tissue samples, can be analyzed on a single “chip”—a small glass surface that carries an array of microscopic points that indicate each molecule or sample that is being studied.

DNA Microarrays
DNA microarrays can be used to analyze an entire genome on one chip. This provides a whole picture of genetic function for a cell or organism, rather than a gene-by-gene approach.
Scientists can use DNA microarrays to:
●● detect mutations in disease-related genes.
●●monitor gene expression.
●● diagnose infectious diseases and identify the best antibiotic treatment.
●● identify genes important to crop productivity.
●● improve screening for microbes used in environmental cleanup.

DNA-based arrays are essential for using the raw genetic data provided by the Human Genome Project and other genome projects to create useful products. However, gene sequence and
mapping data mean little until we determine what those genes do—which is where protein microarrays can help.

Protein Microarrays
The structures and functions of proteins are often much more complicated than those of DNA, and proteins are less stable than DNA. Each cell type contains thousands of different proteins,
some of which are unique to that cell’s job. In addition, a cell’s protein profile—its proteome varies with its health, age, and current and past environmental conditions.
Protein microarrays may be used to:

●● discover protein biomarkers that indicate disease stages.
●● assess potential efficacy and toxicity of drugs before clinical trials.
●●measure differential protein production across cell types and
developmental stages, and in both healthy and diseased states.
●● study the relationship between protein interactions and function.
●● evaluate binding interactions between proteins and other molecules.

The availability of microarray technology has enabled researchers to create many types of microarrays to answer scientific questions and discover new products.

Tissue Microarrays
Tissue microarrays, which allow the analysis of thousands of tissue samples on a single slide, are being used to detect molecular profiles in healthy and diseased tissues and validate potential drug targets. For example, brain tissue samples arrayed on slides connected to electrodes allow researchers to measure the electrical activity of nerve cells exposed to certain drugs.

Whole-Cell Microarrays
Whole-cell microarrays alleviate the problem of protein instability in microarrays and permit a more accurate analysis of protein interactions within a cell.