D.N.A. Technology and Recombinant Technologies | Anthropology Optional for UPSC PDF Download

Introduction

• Recombinant DNA technology, also known as genetic engineering, is a method used to create artificial DNA by combining genetic materials from different sources. This technique was developed following the discovery of restriction enzymes in 1968 by Swiss microbiologist Werner Arber.
• Creating recombinant DNA and introducing it into a host organism is a complex process. First, the desired gene must be identified and selected. Then, a suitable vector must be chosen to integrate the gene and form the recombinant DNA.
• Once the recombinant DNA is created, it must be introduced into the host organism. Finally, the recombinant DNA must be maintained within the host and passed on to its offspring.

Tools of Recombinant DNA Technology

• Enzymes play a crucial role in recombinant DNA technology, with restriction enzymes cutting DNA strands, polymerases synthesizing new strands, and ligases binding them together. Restriction enzymes, which can be endonucleases or exonucleases, are particularly important in determining where the desired gene is inserted into the vector genome. Endonucleases cut within the DNA strand, while exonucleases remove nucleotides from the ends of strands.
• Restriction endonucleases are sequence-specific, usually recognizing palindrome sequences, and cut DNA at specific points known as restriction sites. This creates sticky ends in the sequence. To ensure complementary sticky ends, both the desired genes and the vectors are cut with the same restriction enzymes, making it easier for the ligases to bind the desired gene to the vector.
• Vectors play a vital role in transporting and integrating the desired gene into the host organism. Plasmids and bacteriophages are common vectors in recombinant DNA technology due to their high copy number. Vectors consist of an origin of replication (a nucleotide sequence where replication begins), a selectable marker (genes providing antibiotic resistance, such as ampicillin), and cloning sites (sites recognized by restriction enzymes where desired DNA is inserted).
• The host organism is the final tool in recombinant DNA technology, responsible for accepting the engineered vector containing the desired DNA. Several methods exist for inserting recombinant DNA into the host, including microinjection, biolistics (gene gun), alternate cooling and heating, and the use of calcium ions.

Process of Recombinant DNA Technology

The entire process of recombinant DNA technology involves several steps that must be carried out in a specific order to produce the desired product.

Step 1: Isolation of Genetic Material

• The initial step in recombinant DNA technology is isolating the desired DNA in its purest form, meaning it should be free from other macromolecules.

Step 2: Cutting the Gene at Recognition Sites

• Restriction enzymes play a significant role in determining the location where the desired gene is inserted into the vector genome. These reactions are known as "restriction enzyme digestions."

Step 3: Amplification of Gene Copies through Polymerase Chain Reaction (PCR)

• Once the target gene has been cut using restriction enzymes, PCR is used to amplify a single copy of the DNA into thousands or even millions of copies.

Step 4: Ligation of DNA Molecules

• During the ligation step, the two pieces of DNA – the cut fragment and the vector – are joined together with the help of the enzyme DNA ligase.

Step 5: Insertion of Recombinant DNA into the Host

• In this step, the recombinant DNA is introduced into a recipient host cell, a process known as transformation. Once the recombinant DNA is inserted into the host cell, it multiplies and is expressed in the form of the manufactured protein under optimal conditions.

As noted in the tools of recombinant DNA technology, there are various methods for achieving this transformation. The successfully transformed cells or organisms pass on the recombinant gene to their offspring.

Application of Recombinant DNA Technology

• Recombinant DNA technology is a powerful tool used in various fields, including agriculture, medicine, and research. It involves the manipulation and combination of DNA from different sources to create genetically modified organisms (GMOs) or produce specific proteins.
• One application of recombinant DNA technology is in the detection of HIV in a person. Gene therapy is another application, which involves correcting genetic defects that cause hereditary diseases. In clinical diagnosis, recombinant DNA technology is used in techniques like ELISA to detect the presence of specific antigens or antibodies.
• In agriculture, recombinant DNA technology has been used to create GMOs, such as Flavr Savr tomatoes with improved shelf life, golden rice enriched with proteins, and Bt-cotton, which is resistant to pests like bollworms.
• In medicine, recombinant DNA technology has been instrumental in the production of insulin, a vital hormone for people with diabetes.

DNA Cloning

• DNA cloning is a process that generates multiple copies of a specific DNA fragment or gene, creating identical copies called a DNA clone. This is accomplished by inserting the desired DNA fragment into a small DNA molecule capable of replicating within a living cell, such as a bacterium. This small replicating molecule is referred to as a DNA vector.
• Commonly used vectors include yeast cells, viruses, and plasmids. Plasmids are circular DNA molecules derived from bacteria that are separate from the main cellular genome. They carry genes that provide the host cell with beneficial properties, such as mating ability and drug resistance. Plasmids are convenient for DNA cloning because they are small and can carry additional DNA that can be incorporated into them.

Applications of  Gene Cloning

• Gene cloning has a variety of applications in different fields, including medicine, agriculture, and scientific research.
• In the medical field, gene cloning plays a crucial role in the production of essential biological substances such as hormones, vitamins, and antibiotics. These substances can be used for various treatments and therapies, improving the health and well-being of individuals.
• In agriculture, gene cloning can be employed to enhance crop productivity and improve plant health. For example, cyanobacteria are capable of nitrogen fixation, a process that can be optimized using specific genes. By incorporating these genes into crops, farmers can reduce their reliance on chemical fertilizers, leading to more sustainable and eco-friendly agricultural practices.
• Gene cloning is also an essential tool in scientific research, as it allows scientists to identify and manipulate specific genes. By isolating a particular gene and growing it in a controlled environment, researchers can better understand its functions and potential applications.
• Lastly, gene cloning plays a significant role in gene therapy, where a malfunctioning gene is replaced with a healthy one. This technique has the potential to treat various genetic disorders, such as leukemia and sickle cell anemia, by correcting the underlying genetic cause.

Conclusion

Recombinant DNA technology and gene cloning are powerful tools that have revolutionized various fields, including agriculture, medicine, and research. These techniques involve the manipulation and combination of genetic material from different sources to create genetically modified organisms or produce specific proteins. Applications of recombinant DNA technology include the detection and treatment of diseases, the production of essential biological substances, and the enhancement of crop productivity. Gene cloning enables the identification and manipulation of specific genes for various purposes, including gene therapy and scientific research. Overall, these advances in genetic engineering hold great promise for improving human health, developing sustainable agricultural practices, and furthering our understanding of the genetic basis of life.