Recombinant DNA Technology

The aim of recombinant DNA technology involves cloning a specific gene. It is possible to produce multiple identical copies of a gene, a molecule, a cell through gene cloning. Recombinant DNA technology involves synergizing genetic material from different sources to create a new gene or a cell. Genetic engineering plays an important role in determining the structure of a gene, detecting carriers, diagnosing genetic disorders and gene therapy. Many hormones such as insulin, glucagon, and growth factors get synthesized using genetic engineering and used for commercial purposes. Before the advent of genetic engineering, there were limited sources for improvising crop quality, yield, agricultural techniques, and plant breeding.

However, with the recombinant DNA technology or genetic engineering, the yield and the crop quality started improving. Introduction of Stress and drought tolerance genes turned out to be successful. BT-Cotton is a classical example of recombinant DNA technology. It is a genetically modified crop involving resistance to bollworm infection. The plant gets modified with a specific gene helping the plant from bollworm attack. The gene product is a form of a protein. It gets released in the plant as soon as the worm attacks the plant. The worm immediately dies when it comes in contact with the gene product. Thus, BT-Cotton helped the farmers and the cotton industry grow. Many such success stories heard in the world involve the hard work of the recombinant DNA technologists. A body of techniques in recombinant DNA technology includes cutting the genes apart and splicing them with different pieces of DNA.

Image: Recombinant DNA technology

Materials used for recombinant DNA technology:

Recombinant DNA technology uses materials such as cells, bacteria, viruses, pieces of DNA, restriction digests, chromosomes, plasmid vectors, and many other substances. Commonly used organisms include bacteria as host organisms since they are single-celled structures. These bacteria are genetically engineered to produce vaccines, hormones, enzymes, and other biomolecules. For example, E. coli bacteria are genetically engineered to synthesize hormone insulin and growth factors. They also synthesize plant metabolites. An autonomously replicating DNA or a plasmid mostly depicts a circular, double-stranded DNA. Plasmids used in recombinant DNA technology replicate at each cell division. They are relatively convenient in isolation. Recombinant DNA technology allows the modification and manipulation of genes within or between the species. Hence, modification of plasmids involves restriction enzymes cleaving at a specific site and allowing foreign DNA inserts.

Recombinant DNA technology also utilizes viral vectors. However, it requires host genetic machinery for its replication.

Steps involved in recombinant DNA technology:

Cloning generates a series of DNA fragments. The first step involves cleaving the DNA segment at specific sites with the restriction enzymes. Restriction enzymes recognize and cut short sequences. They create a staggered or a blunt end in the DNA double helix. Now comes the role of a vector. A vector such as a virus or a plasmid carries a desired gene into the host and produces multiple copies.

Recombination:

Restriction enzymes cleave the DNA and produce either staggered or sticky ends. Thus, the plasmid DNA combines with foreign DNA. Their ends get sealed and stabilized through the action of another enzyme known as DNA ligase. The product obtained out of this activity is known as recombinant DNA. 

Transfer of the recombinant vector to the host:

The recombinant DNA molecules get introduced into the host organism. Once the recombinant plasmids get transformed into the host, they start multiplying themselves. The process develops identical foreign DNA molecules known as clones. A plasmid consists of genes showing resistance to antibiotics. The genes help the host to save themselves from the action of antibiotics. The absence of antibiotic resistance genes makes the bacteria sensitive to antibiotics. Screening of the clones involves nucleic acid hybridization.

References:
[1] Recombinant DNA Technology, Sardul Singh Sandhu
[2] Biotechnology-4: Including Recombinant DNA Technology, S. Mahesh

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