Understanding Recombinant DNA
How Gene Splicing Can Produce Unique Nucleotide Sequences
Jul 24, 2008
Recombinant DNA is commonly used to manufacture therapeutic agents such as insulin, an important hormone administered to diabetic patients. The genetic sequence most commonly synthesized by scientists includes fragments of DNA from several different organisms and is sometimes referred to as “chimeric” DNA, after the mythological chimeras that existed as a blend of several different animals.
There are three major techniques scientists use to manufacture recombinant DNA: transformation, phage introduction, and non-bacterial transformation. Though each method has its nuances, they all have the same basic procedure as transformation.
Transformation
Transformation begins after the scientist has identified the piece of DNA from a particular genome she wants to copy for future pasting. A restriction enzyme must then be employed to physically cut the desired sequence. Restriction enzymes (or restriction endonucleases) are naturally occurring enzymes produced by bacteria to protect against viral infection. While inside its host, a restriction enzyme will cut up foreign DNA, but ignore the host genome which has been marked via methylation.
Hundreds of restriction enzymes have been identified and isolated, each specific for a particular sequence, giving the scientist adequate selectivity when selecting sequences for integration. After the desired portion of the genome has been cut from one organism, it is then incorporated into a vector, such as a virus or a plasmid, using DNA ligase. The vector can be thought of as a vehicle whose primary purpose is to shuttle DNA from one organism to another.
In order for one strand of DNA to be incorporated into another, both strands must be treated with the same restriction enzyme. After the enzyme does its work, it leaves behind jagged DNA molecules with overhanging pieces called “sticky ends.” The sticky ends can be glued together with the aid of another enzyme, DNA ligase.
Often, an antibiotic marker is included with the vector so that the scientist can identify host cells that have taken up the foreign genetic material. If an antibiotic agent is administered to a bacteria culture, those without the recombinant DNA will selectively die out, leaving behind the hosts who have the resistance-marker (and thus the vector) inside them.
Uses of Recombinant DNA
Recombinant DNA is most commonly known for its service in medicine, particularly for the production of synthetic insulin. By incorporating the human gene for insulin into bacterial plasmids (which reproduce quickly, and in great numbers), this essential regulator of blood glucose can be produced in the lab, bottled, and then administered to patients with diabetes. Recombinant DNA is also used in agriculture to make crops that produce their own pesticides, or are better able to cope with extreme climate.
References
Alberts, Bruce, Karen Hopkin, Alexander Johnson, Julian Lewis, Martin Raff, Keith Roberts and Dennis Bray. Essential Cell Biology. 2nd ed. New York: Garland Science, 2004, pp. 328; 352-360.
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