Why Are Bacteria Used in Recombinant DNA Technology?
The first question to ask yourself is: why are bacteria used in recombination technology? Bacteria, plasmids, viruses, and plant cells are all viable options. However, a question of ethicality needs to be asked: should bacteria be used at all? If so, what are the risks and benefits? I’ll answer this question below. In the meantime, read up on this technology by Griffiths, Anthony JF.
Escherichia coli
Recombinant DNA technology is a process in which a bacterial strain is transformed with a gene of interest. This gene is transcribed into a protein by a multicomponent system. This complex assembly comprises three distinct regions: the promoter, a -35 box, and a spacer region. The promoter of E. coli is highly similar to the consensus promoter sequence obtained by aligning many different plasmids. The consensus promoter sequence, which is representative of the best promoter sequence, has one or two deviations in the -35 box. Furthermore, the recombinant protein must be able to bind to the cell membrane or to other structures to effectively stop the process of transcription.
The expression of recombinant gene products in E. coli is affected by the bacterial host’s characteristic temperature. Lower temperatures, such as 37degC, promote inclusion of body formation and inhibit bacterial growth, while higher temperatures promote protease activity. Further, lower temperatures may require longer induction periods to produce sufficient amounts of the recombinant protein.
In the laboratory, a mixture of transformed bacterial cells is spread on the growth medium in a Petri dish. These cells are invisible to the naked eye, but as they undergo successive rounds of cell division, colonies form. These colonies are known as DNA clones. The clones produced in one dish represent hundreds of different DNA fragments, called clones. The colonies can be grouped together and stored in a DNA library.
Recombinant DNA technology also improves b-lactam antibiotic producers. In the commercial sector, recombinant DNA technology enables the creation of strains with more than one b-lactam gene. The bacteria used in this process are commonly called’recombinagenic’, but they are often referred to as ‘transgenic’ strains.
plasmids
Plasmids are a type of genetic material with many copies of DNA that contains genes that can be used to create useful proteins. For example, the human insulin gene is contained within a plasmid that can be expressed in E. coli bacteria to produce insulin. The plasmid is then treated with restriction enzymes and DNA ligase to remove unwanted sequences. Bacteria that take up the plasmid can be identified by antibiotic selection.
During the process of recombinant DNA technology, scientists use a tool known as a plasmid to make genetically modified proteins. The plasmids are circular DNA molecules that originate from yeast cells or viruses. They carry genes that control the properties of their host cells, such as mating ability and toxin production. In addition, these plasmids are small enough to be handled easily.
In addition to gene therapy, plasmids are commonly used in gene transfer and genetic engineering experiments. These tiny replicons are capable of independent replication when inserted into the correct host. They can also be used to create GMO and GMP plants. Similarly, plasmids are used in gene mapping and cloning experiments. Further, recombinant DNA technology enables the creation of new, genetically modified organisms, which are better able to resist antibiotics.
This technique is also used for creating clones. These clones are created by inserting a gene into a circular piece of DNA called a plasmid. The clones are genetically identical, and the plasmids contain the gene that was intended to be expressed. In addition to creating clones, recombinant DNA technology allows scientists to produce many copies of a single gene.
Viruses
Viruses are used in recombined DNA technology to produce proteins that are not found in their natural form. In addition to making vaccines, these viruses are used as gene therapy vectors. They have the potential to transform plants, and some scientists believe that they may even be able to create human proteins. But, for the time being, recombinant DNA technology is still quite experimental.
The first step in this process is to introduce transgenes into the recombinant virus. This is done by inserting the desired gene into a shuttle vector or pAdTrack-CMV. This DNA molecule is linearized by a restriction endonuclease, then transformed into an adenovirus supercoiled vector. The recombinant adenovirus is then transfected into a packaging cell line.
The first animal cells were created about a decade after Berg’s team. Most early studies used mouse cells, and the researchers introduced rabbit DNA fragments into murine germ-line cells. They had already established that foreign genes could be integrated into somatic cells, but this was the first time they had done it with germ cells. This breakthrough made recombinant DNA a widely-accepted technique.
Another advantage of using virus genes is their ability to help plants fight disease. Researchers have been able to use DNA technology to increase the efficiency of plant growth by modifying the genes of bacteria that fix nitrogen in the soil. The plants can now use nitrogen from the atmosphere without having to rely on bacteria to produce proteins. This has allowed scientists to create new plants that are resistant to viral diseases. It’s a win-win situation for the environment and humankind.
Plant cells
In addition to being a sustainable alternative to animal cell cultures, plants are also a good option for producing functional recombinant proteins. Because plants produce proteins with a variety of structural characteristics, they are also highly cost-efficient. Plant-derived recombinant proteins may even be of benefit to developing countries, as they can be produced at a low cost. Here’s why.
Recombinant DNA methods are used in drug discovery and development, as they are commonly used to isolate and improve specific proteins. Additionally, they help test the potential effects of therapeutic molecules on cell physiology and toxicity. Recombinant methods are also used to optimize natural product therapeutics, such as increasing yields or adjusting chemical structures. Ultimately, this technology helps improve plant health and yields.
One of the major advantages of transient expression over transgenic plants is that yield is higher. The improved transgene delivery is attributed to the elimination of the “position effect”. Earlier gene delivery methods consisted of soaking leaves in Agrobacterium culture, and only the outer layer of the leaf could be exposed to the transgene. This technique is highly inefficient, however, and limits scalability.
Indirect transgene delivery uses the natural ability of plant viruses and some pathogenic agrobacteria species to transfer genomes and tumor-inducing plasmid DNA. A plant virus may carry infectious viral particles that allow the transgene to enter plant cells. While these viral particles may be harmful, a plant expression system may be safer than other systems. Agrobacterium T-DNA can reach a very large number of plant cells.
Vaccines
Recombinant DNA technology uses bacteria or yeast cells to produce vaccines. The scientists take a small piece of the target virus’ DNA and insert it into the bacteria or yeast cells used for vaccine production. For example, in the hepatitis B vaccine, scientists insert the DNA code for a protein found on the surface of the virus into yeast cells. This purified protein then acts as the active ingredient in the vaccine.
These vaccines are produced using circular double-stranded DNA plasmids. They contain encoded antigens that cause a cellular response by activating cytotoxic T lymphocytes. DNA vaccines need to be adjuvanted to be effective. A naked DNA fragment will not work like a vaccine, so they need to be packaged in a biodegradable carrier.
DNA vaccines are currently used in veterinary medicine, especially for melanoma in dogs. Clinical trials for these vaccines are underway in the U.S., where the number of patients for these vaccines is unmanageable. However, the delivery of DNA-based antigens to the body is the biggest challenge. DNA vaccines must penetrate the cell membrane and reach the nucleus, where the replication machinery resides. In addition, a needle will not reach the nucleus.
RNA vaccines are also in development. Unlike DNA vaccines, RNA vaccines do not provide the protein antigen to the body. Instead, they supply the genetic instructions needed for the antigen to be produced by cells within the body. When these cells produce the antigen, the immune system reacts and produces an antibody against the infection. This method is relatively simple and quick, and it holds great promise for vaccine development in the future.