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Recombinant DNA

life science Maturity 9-11 Next article →

Recombinant DNA, often called rDNA, consists of DNA molecules created through laboratory methods of genetic recombination. These methods bring together genetic material from multiple different sources. This process creates new sequences that would not naturally exist within a single genome. Because these molecules can be made from two different species, they are sometimes called chimeric DNA. This name refers to the mythical chimera, a creature composed of parts from different animals. Recombinant DNA is possible because all organisms share the same basic chemical structure. The only difference between species is the specific sequence of their nucleotides.

recombinant formation of plasmids
recombinant formation of plasmids

Scientists use a process called molecular cloning to produce these molecules. Molecular cloning is one of two main methods used to direct the replication of specific DNA sequences. The other common method is called polymerase chain reaction, or PCR. There are two fundamental differences between these techniques. PCR replicates DNA inside a test tube without the use of living cells. In contrast, molecular cloning involves replicating DNA within a living host cell. While PCR amplifies an existing sequence, cloning requires cutting and pasting different DNA sequences together.

To perform molecular cloning, researchers must use a cloning vector. A vector is a small DNA molecule that can replicate inside a living cell. These vectors are generally derived from viruses or plasmids. They contain the necessary genetic signals for replication. Vectors also include extra elements to help insert foreign DNA or identify cells that contain the recombinant DNA. The choice of vector depends on the host organism and the size of the DNA being cloned. The standard cloning protocol involves seven specific steps. First, a host organism and vector are chosen. Then, the vector DNA and the target DNA are prepared. Next, the recombinant DNA is created and introduced into the host. Finally, scientists select and screen the organisms to find the desired clones.

Once the recombinant DNA is inside a host, it may undergo DNA expression. This is the process where the cell uses the DNA to produce a recombinant protein. To achieve this, scientists must perform transfection, which is the introduction of DNA into host cells. Common host cells include bacteria, yeast, insect, or mammalian cells. Scientists often use human embryonic kidney cells or Chinese hamster ovary cells. Simply putting DNA into a cell does not guarantee expression. The DNA might be replicated without ever being used to make a protein. To ensure expression, the gene often requires restructuring. It must include parts like a promoter, a translational initiation signal, and a transcriptional terminator. These parts allow the host's machinery to read the foreign instructions.

Most organisms containing recombinant DNA maintain a normal phenotype. This means their appearance, behavior, and metabolism usually stay the same. Scientists often use a polymerase chain reaction (PCR) test to detect the presence of the new sequences. However, some changes can occur. The recombinant gene might cause toxicity to the host organism. It can also cause insertional inactivation. This happens when the recombinant DNA inserts itself directly into a host cell's gene. Researchers sometimes use this to "knock out" a gene to study its function. In other cases, the insertion might inappropriately activate a previously silent host gene.

Gene cloning
Gene cloning

Recombinant DNA technology has massive applications in medicine and industry. It is used in basic research to identify, map, and sequence genes. It is also used to create recombinant proteins, which serve as reagents in many laboratory experiments. One famous example is recombinant chymosin. Chymosin is an enzyme used to make cheese. Traditionally, it was taken from the stomachs of calves. Now, scientists use engineered E. coli bacteria to produce it. This microbiologically produced enzyme is identical to the calf version. Today, about 60% of hard cheese in the United States is made using this genetically engineered chymosin. The FDA granted it "generally recognized as safe" status in 1990.

Medicine has also been transformed by this technology. Recombinant human insulin has almost entirely replaced insulin taken from pigs or cattle. This is vital for patients with type 1 diabetes. Using recombinant insulin prevents the immune system from attacking the medicine. Similarly, recombinant human growth hormone (HGH) is used for patients with growth deficiencies. Before this, HGH was taken from human cadavers, which was an unsafe practice. Recombinant factor VIII is also used to treat hemophilia. This protein helps blood clot in patients who cannot do so naturally. Finally, the hepatitis B vaccine is created using a recombinant subunit method. This involves producing a virus surface antigen within yeast cells.

recombinant formation of plasmids
recombinant formation of plasmids
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