Adenoviruses are clusters of DNA that contain viruses. These DNA groups trigger respiratory disease, and they even cause people to develop one form of the common cold. However, medical researchers have discovered that they can use these DNA clusters in gene therapy to treat cystic fibrosis as well as other diseases. To use them for treatments, medical professionals must modify an adenovirus with a particular type of tumor fighting gene. We have outlined the adenovirus infection protocol in this article.
Cell modifications and transfers are completed through a scientific process called transduction. The process involves conveying or introducing virus-containing DNA clusters or a foreign gene into a separate cell. To make the transfer, researchers need a viral vector, which is a tool used to transmit the DNA into a cell. Adenoviruses, retroviruses and lentiviruses are three types of viral vectors. With transduction, science professionals can infect most cells without affecting their ability to grow and develop. Transfection is another DNA transferring process. Scientists select their transferring methods based on the type of genetic experiments they are conducting during the adenovirus infection protocol.
Adenovirus Infection Protocol Background
Many years ago, scientists discovered the helpful properties of adenoviridae. In fact, since the virus-containing DNA clusters have an effect on cells that replicate and cells that do not, medical professionals often use them for gene therapy treatments and follow an adenovirus infection protocol. The ability to contain large transgenes as well as hold protein codes without needing to be incorporated into an organism’s genetic material are other benefits of using recombinant adenovirus.
Recombinant adenoviruses give researchers a versatile way to process gene expression experiments and complete therapeutic functions. Medical professionals use recombinant adenoviruses to transfer genes in vitro, during gene therapy procedures and for in vivo vaccinations. In most cases, researchers use one of two techniques to generate recombinant adenoviruses. The first method consists of connecting adenoviral genome DNA pieces to restriction endonuclease portions that feature a transgene. However, this technique is challenging for a research team because of the decreased effectiveness of large fragment connections as well as a lack of distinctive restriction sites.
Most medical researchers select the second method, which involves the use of similarly functioning gene combinations that perform comparably in mammalian cells. In addition, the mammalian cells must be able to complement faulty adenoviruses. When similarly functioning gene combinations are united, the result is a defective adenovirus that has the ability to duplicate in the packaging line to provide the absent gene products. Researchers identify the genetically engineered DNA by analyzing particular plaques that are produced in the bacteria layer of the packaging cells. Despite the practicality of this method, a number of disadvantages come with it. For instance, uniting similarly functioning gene combinations is an inefficient process. In addition, researchers must repeat several rounds of the plaque purification process, and the completion period of the viral making process is lengthy. These issues have prevented medical teams from advancing adenoviral vector technology. However, recombinant adenovirus is still widely considered the most popular vector in genomics research because it has a wide host range of mammals, and the virus remains epichromosomal after it has been introduced into the cell (it does not integrate itself into the host chromosome).
Adenovirus Infection Protocol Process
To modify adenoviruses, a general adenovirus infection protocol is followed. Start the process one day earlier than the infection day. To do this, set up a six well plate so that it is about 50 percent convergent for the day of the infection. Before starting the actual modification process, make sure that the cells have come together. If the cells are ready, warm the virus with ice. Keep in mind that viruses must be stored at -80 degrees Celsius when they are not being used. If they are stored at cooler temperatures, they may become less concentrated. The next step is to add a particular Multiplicity of Infection, or MOI, of the virus to every well in the plate based on the selected dose curve. During the adenovirus infection protocol, be sure to have a zero MOI for a negative control to check cell growth and death. To determine the proper virus amount for a particular MOI, take the number of cells and multiply it by the desired MOI. This formula will equal the total Plague Forming Units, or PFUs, needed. Be sure to add the right virus amount to each dose well. Also, label the wells with the MOI used. The next step is to deposit the virus-infected cells in an incubator that is set at 37 degrees Celsius. Allow the cells to incubate for 24 hours. You are now ready to begin your research using the adenovirus infection protocol outlined above.