25/05/2024 11:57 PM

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The Health Maniacs

What are the uses of plasmid?

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Plasmid

A plasmid is a small, circular, occasionally linear, self-replicating unit of DNA that is not part of the chromosome. Few genes are included on each plasmid. Several techniques rely on plasmid DNA as an input, including transfection, sequencing, clone screening, restriction digestion, cloning, and polymerase chain reaction. There are several different ways to extract plasmid DNA from bacteria, and both are effective. The alkaline lysis technique is by far the most popular method used. Commercially available plasmid DNA purification kits today ensure the highest possible quality and purity in minutes. Miniprep, midiprep, maxiprep, etc., are all different types of plasmid preparation distinguished by the size of the bacterial culture used (and thus the plasmid DNA yield). Minipreparations of plasmid DNA is frequently used to check bacterial clones for recombinant DNA inserts. Here, we introduce a quick and easy protocol developed by Holmes and Quigley (1981) that can be used to screen numerous colonies or small cultures for recombinant DNA inserts. After collecting bacterial cells, they are briefly boiled to inactivate any toxins or pathogens and then centrifuged to separate the genomic DNA from any remaining cellular debris. Plasmids are extracted using isopropanol precipitation followed by RNase treatment. This method successfully produces sufficient plasmid DNA for multiple restriction digests, and it applies to plasmids with sizes ranging from 3.9 to 13.4 kb. It’s possible to process several samples simultaneously.

Importance

In biotechnology, plasmids serve various functions and occur in multiple sizes. In the 1970s, they were used to introduce genes into bacteria to stimulate their manufacture of therapeutic proteins like human insulin. This was a significant breakthrough in the field of recombinant DNA.

More recently, plasmid DNA has been studied as a potential therapeutic platform for treating viral, genetic, and acquired illnesses. For instance, plasmid DNA is useful for creating DNA vaccines against infectious diseases such as HIV/AIDS, Ebola, Malaria, gastrointestinal infections, and influenza. To immunize against a disease, the plasmid must be changed genetically to generate just one or two of the target proteins. Compared to older immunization methods, DNA vaccines have several benefits. First, they prevent you from having to inject harmful substances. Second, they elicit a reaction from the immune system’s B and T cells. Thirdly, they are more convenient to store and move since they are less susceptible to temperature changes. In conclusion, they are cheap and easy to produce in significant quantities.

There are substantial obstacles to using plasmids in the vaccine development process. There has been an insufficient human immune response to most DNA vaccines tried in clinical trials for them to be effective in preventing illness. However, in 2006, promising findings were reported from early clinical trials of a DNA vaccine intended to combat H5N1 avian flu. Ischemic stroke, Parkinson’s disease, Alzheimer’s disease, and multiple sclerosis are currently incurable neurological diseases being researched as potential DNA vaccine targets. In 2007, researchers conducted early research on using DNA immunization to treat multiple sclerosis. However, further testing is required to verify the efficacy of DNA plasmids and their application as DNA vaccines.

Discovery

In the late 1940s, scientists looking into the mechanisms by which bacteria develop resistance to antibiotics and how phages (bacteria-specific viruses) and other DNA structures can transmit traits to their offspring discovered that bacterial cells contained DNA strands that were structurally distinct from those of chromosomes. At various points throughout history, scientists have referred to these DNA molecules as pangenes, bioblasts, plasmalogens, plastogenes, choncriogenes, cytogenetics, proviruses, and episomes.

First used in context by Joshua Lederberg in 1952, the term “plasmid” has since entered the English language. “any extrachromosomal hereditary element,” in his words. Lederberg coined the term “P22 jacket” in a report on his work with Salmonella bacteria and his doctoral student Norton Zinder’s trials with the P22 virus. Transduction refers to the process whereby viral particles acquire genes from one host organism and transmit them to another. For a long time, though, nobody knew why this was happening or how it happened. This shifted with the 1953 discovery of the DNA double helix shape, which established beyond a reasonable doubt that genetic material constitutes DNA. Scientists quickly learned that plasmids were made up of short DNA stretches that allowed for certain features to be transmitted.

In the 1960s, researchers had already discovered several different plasmids. Esther Lederberg, Joshua’s wife, was the first to discover fertility plasmids in the late 1940s; these plasmids carry the genes for fertile bacterial conjugation. Plasmids carrying genes for antibiotic or toxin resistance are referred to as “resistance” (R) plasmids. The first molecule of recombinant DNA was created with the help of an R plasmid (pSC101).

Application

Plasmids, which may be found in bacteria, archaea (single-celled creatures), and eukarya, are DNA strands or loops that carry genetic material (organisms of complex cell structure). While chromosomes are the significant structures in cells that hold DNA, plasmids are more diminutive, self-contained DNA units that can carry only a few genes. Plasmids are self-replicating DNA elements that bacteria from their surroundings may take up and pass along to other species.

The host organism relies on plasmids as a means of enduring environmental stress. Numerous plasmids, for instance, include the genetic instructions to manufacture antibiotic or toxin-degrading enzymes. Some have genes that aid the host organism in processing foreign chemicals or eliminating harmful germs.

Plasmids are convenient for genetic engineering because of their adaptability and other features. They have shorter DNA sequences than other organisms, ranging from 1,000 to 20,000 base pairs. Second, they snap back into shape after being sliced open without disintegrating. Plasmids may now have fresh DNA inserted into them with relative ease. To generate an infinite supply of copies after a DNA change, the changed plasmid can be cultured in bacteria and subjected to self-replication.

Plasmids are essential in genetics and biotechnology labs due to their extended shelf life and ease of modification. Most prominently, they serve as a vector to transport foreign DNA into bacteria, an essential procedure for genetic engineering and other biotech applications.

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